Road Rage Among the Polymerases

DNA polymerase is faster than RNA polymerase. RNA polymerase also leaves detritus in its wake. What happens when they collide?

DNA is a country road- one lane, two directions. Yet in our cells it can be extremely busy, with transcription (RNA synthesis) happening all the time, and innumerable proteins hanging on as signposts, chemical modifications, and even RNA hybridized into sections, creating separated DNA structures called R-loops. When it is time for DNA replication, what happens when all these things collide? One might think that biology had worked all this out by now, but these collisions can be quite dangerous, sending the RNA polymerase careering into the other (new) DNA strand, causing the DNA polymerase to stall or miss sections, and causing DNA breaks, which activate loud cellular alarm bells and mutations.

Despite decades of work, this area of biology is still not yet very well understood, since the conditions are difficult to reproduce and study. So I can only give a few hints of what is going from current work in the field. A couple of decades ago, a classic experiment showed that in bacteria, DNA polymerases can be stopped cold by a collision with an RNA polymerase going in the opposite direction. However, this stall is alleviated by a DNA helicase enzyme, which can pry apart the DNA strands and anything attached, and the DNA replication complex sails through, after a pause of a couple of seconds. The RNA polymerase, meanwhile, is not thrown off completely, but switches its template from the complementary strand it was using previously to the newly synthesized DNA strand just made by the passing DNA polymerase. This was an amazing result, since the elongating RNA polymerase is a rather tightly attached complex. But here, it jumps ship to the new DNA strand, even though the old DNA strand remains present, and will shortly be replicated by the lagging strand DNA polymerase complex.

General schematic of encounters between replication forks and RNA polymerases (pink, RNAP). Only co-directional, not head-on, collisions are shown here. Ribosomes (yellow) in bacteria operate directly on the nascent mRNA, and can helpfully nudge the RNA polymerase along. In this scheme, DNA damage happens after the nascent RNA is used as a primer by a new DNA polymerase (bottom), which will require special repair. 

The ability of the RNA polymerase to switch template strands, along with the nascent RNA it was making, suggests very intriguing flexibility in the system. Indeed, DNA polymerases that come up from behind the RNA polymerase (using the same strand as their template) have a much easier time of it, passing with hardly a pause, and only temporarily displacing the RNA polymerase. But things are different when the RNA polymerase has just found an error and has back-tracked to fix it. Then, the DNA polymerase complex is seriously impeded. It may even use the nascent RNA hanging off the polymerase and hybridized to the local DNA as a primer to continue synthesis, after it has bumped off the RNA polymerase that made it. This leads in turn to difficulties in repair and double strand breaks in that DNA, which is the worst kind of mutation. 

The presence of RNA in the mix, in the form of single strands of RNA hybridized to one of the DNA strands, (that is, R-loops), turns out to be a serious problem. These can arise either from nascent transcription, as above, or from hybridization of non-coding RNAs that are increasingly recognized as significant gene regulators. RNA forms a slightly stronger hybrid with DNA than DNA itself does, in fact. Such R-loops (displacing one DNA strand) are quite common over active genomes, and apparently present a block to replication complexes. One would think that such fork complexes would be supplied with the kinds of helicases that could easily plow through such structures, but that is not quite the case. R-loops cause replication complex stalling, and can invoke DNA damage responses, for reasons that are not entirely clear yet. 

A recent paper that piqued my interest in all this studied an ATPase motor protein that occurs at stalled replication forks and helps them restart, presumably by acting as a DNA or RNA pump of some kind, and forcing the replication complex through obstructions. It is named WRNIP1, for WRN interacting protein, for it also interacts with Werner syndrome protein, another interesting protein at the replication fork. This is another ATPase that is a helicase and also a backwards 3' -> 5' exonuclease that cleans up DNA ends around DNA repair sites, helping to remove mismatched and damaged DNA so the repair can be as accurate as possible. As one can guess, mutations in this gene cause Werner Syndrome, a striking progeria syndrome of early aging and susceptibility to cancer. 

While the details of R-loop toxicity and repair are still being worked out, it is fascinating that such conflicts still exist after several billion years to figure them out. It is apparent that the design of DNA, while exceedingly elegant, results in intrinsic conflicts between expression and replication that are resolved amicably most of the time. But when either process gets overly congested, or encounters unexpected roadblocks, then tempers can flare, and an enormous apparatus of DNA damage signaling and repair is called in, sirens blaring, to do what it can to cut through the mess.

• Who really believes in climate change?
• The very strong people of the GOP. 
• The ancient Easter Islanders mixed with South Americans.

Jimmy Carter, on Work

Jimmy Carter's "An Hour Before Daylight".

One marked contrast between the recent political conventions was the presence of former presidents. The Republicans had none, (excepting the candidate), not even the very-much alive George W. Bush, or past candidates such as Mitt Romney. The Democrats had two, plus Hillary Clinton, not to mention the current president, Joe Biden. There was additionally a representative of a fourth, Jimmy Carter, to say that he will be happily voting for Kamala Harris in the fall. It speaks to the extremist journey the Republican party has been on, compared to much more conventional (sorry!) path of the Democrats, with recognizably consistent values and respect for character and institutions, both their own and those of the country at large.

None of these Democratic leaders grew up rich. Each was formed in modest circumstances, before joining the meritocracy and working their way up. The Democratic party is now generally viewed as the party of educated people, government workers, and minorities, against the Republican coalition of the very rich and the very poor. One might summarize it as strivers through the educational system, as opposed to strivers through the capitalist system. For one group, being kind, smart, and hard-working are the annointing signs of god, while for the other, it is being rich. Some (usually Republicans) may think these are equivalent, but the current candidates demonstrate the opposite.

This theme is exemplified by the career of Jimmy Carter, who worked his way through Annapolis and a naval career partly spent in the naval nuclear program under Hyman Rickover, then worked his way to the Georgia governorship, the Presidency, and then kept on working through retirement, churning out books and doing good works. The finest of his books, (which are, frankly, a mixed bag), is apparently his memoir of his early life and environment, "An Hour Before Daylight". The theme, for me, was work- hard work. Carter grew up on a large farm, and worked constantly. The book's title comes, naturally, from when the farm day starts. There are pigs to feed, eggs to collect, cows to milk. There are fields to plow, trees to chop down, fences to mend, products to sell, and supplies to buy. The work was evidently endless, as it is on any family farm, and while Carter tells of many swimming, hunting, amorous, and other expeditions, it is the cycle of chores and worries around the farm that was clearly formative.

Jimmy with family, in his Sunday best.

But he was not the hardest worker. His family owned a lot of land, and in this segregated time during the depression, had numerous sharecropping tenants and employees, all black. Carter comments gingerly about this system, balancing his worship of his father with clear descriptions of the hopelessness of the tenant's position. They worked without dreams of attending Annapolis, or inheriting a large estate. Rather, debt was the typical condition, as the Carters ran the supply store as well as owning the land. Carter looked up to many of these employees and tenants, and recounts very close and formative relations throughout his childhood, with both black children and adults. At least until he was drawn, as the system had designed it, into the segregated churches and schools.

Jimmy at his most intense, a naval graduate.

It is hard to grasp, in our heavily urbanized and regulated existence, where work is a 9-5 job and we dream of weekends, family leave, remote work, and retirement, how much work went into a normal existance like this on a farm. Success depended not only on unstinting work, but on an even temper, shrewd foresight, family support, good community relations (including church attendance), and a lot of luck. The wealth and power of the US was built on this kind of scrabbling for economic survival and advancement. The capitalist system continually applied the screws, lowering prices for cotton when too much was being produced, a particular crisis during the depression. Carter tells of the continual inventiveness that his family devoted to new ventures, like selling flavored milks, roasted pecans, sugar cane syrup, boiled peanuts, and tomato catsup, all from their own crops. Not everything was successful, but there was a continual need, even in this out-pf-the-way rural area, to meet the market and keep coming up with new ideas for making money.

Most of all, Carter speaks with pride of his and his family's work. It provided their sustenance, and their relationships, and was thus intrinsically and automatically meaningful. Headed by a benevolent regime, at least as he understood it under his parents, it was an ideal world- busy, endlessly challenging, stimulating, and productive. This is what we need to think about in these end times of the loneliness epidemic and the plague of homelessness and meaninglessness. Religion was a strong presence, but hearing Carter tell it, it weighed relatively lightly on him and his family, (other than sister Ruth, perhaps, who became a renowned evangelist), being more a solace to the poor than a spur to the well-to-do. Their meaning came more from their community and their many and varied occupations. So when people speak of basic income programs, one has to ask whether that really addresses the problem. Much better might be a guaranteed job program, where everyone is offered basic work if they can not find it in the private sector. Productive work that benefits the community, along the lines of the WPA projects of the depression. Work is critical to meaning and mental health, as well as to our communities and nation.

• Zoning and housing.
• Religious nutters lose their minds.
• Another great use of crypto- pig butchering.
• Unbutchering one candidate's garble.
• It smells like the mob.

Wherever Did the Pandemic Go?

Covid has attenuated. But is that from its own evolution, or from our immune reactions to it?

Looking at recent gatherings such as the political conventions and the Olympics, it is evident that the pandemic is over. A graph from the CDC says that mortality from Covid-19 is now similar to influenza- not great, but not catastrophic either, running at roughly a thousand deaths a week, and this with negligible public precautions.

Overall mortality of Covid-19 in the US.

A fundamental scientific and policy question about this is why: did the virus evolve to a less virulent state, or have we evolved (or engineered) enough immunity to fend off the worst? Even after the intense focus on this virus and all the research that has been done, this is a difficult question to answer. There has been a parade of variants, one supposedly more virulent and dangerous than the last, except that we are less affected and increasingly able to ignore them. The scientific community is evidently divided on this causal question, with no good ways to test these basic hypotheses.

I am personally very much in the viral evolution camp, believing that this virus has on its own evolved to be less virulent, even as it gained in transmissibility and ability to evade our immune systems. Surveillance of the virus shows quite high levels this summer, even while its effects are minor, overall. The logic is that this kind of virus does not gain from people shutting themselves up at home and being miserable, let alone dying. Much better for us to be surreptitiously infected and infectious, and able to go about our business, at work and play. We recall that Covid was markedly more lethal at the very outset of the pandemic, before the first set of variants developed. Other cold-type viruses seem to have followed a similar path, and the many zoonotic infections we have picked up (including this one) come from other organisms which carry these pathogens without much difficulty, doubtless after a long evolutionary standoff.

But the graph above makes a different argument, since the vaccines came online around the spring of 2021, reached about fifty percent of the population in late 2021, which is followed by the dramatic drop in covid mortality in spring of 2022. Some researchers point to the lack of attenuation of other pathogens, like HIV, tuberculosis, and smallpox, to say that the evolutionary argument does not hold water. After a pathogen has replicated and spread, (in the case of Covid, in the first week of infection, roughly), it doesn't care what happens to the host- literally whether it lives or dies. They would say that it was the immunization campaign that saved us, and continued infection leading to herd immunity that has created a population increasingly resistant to Covid mortality.

Testing these hypotheses would require Covid-naive populations, which would be ideally split into two study sets, one with vaccination followed by infection, and the other infected directly. This kind of thing may happen as a natural experiment somewhere, and perhaps the closest we can come is the release of Covid restrictions in China. In late 2022/early 2023, China switched abruptly from a zero-tolerance policy of social contact and infection, to a zero-tolerance policy towards bad publicity and accurate mortality reporting, while relaxing anti-Covid restrictions. The result was a surge in death rates, to levels estimated to be higher than those elsewhere, including in the US. This argues that during the restrictive period, the virus had not significantly attenuated via its natural evolution, though then the subsequent mass infection and inoculation did eventually lead in China, as it has elsewhere, to the lower mortality rates seen around the world. 

So, despite the rapidity of viral evolution, one has to conclude that over the short term, the immune hypothesis appears superior to the viral evolution hypothesis, as an explanation of general attenuation of Covid mortality. (Robert Kennedy may disagree, of course!) The evolution of virulence is closely related to the whole lifecycle of a pathogen, especially the way it spreads, making comparisons with other pathogens hazardous. Respiratory pathogens have the opportunity to spread without damaging the host too much, and that seems, in principle, like an advantageous evolutionary path. So I would still hypothesize that over the long term, Covid will settle into a less virulent form that triggers less immune activation (the most lethal aspect of Covid infection), in favor of high transmission and co-existence with our immune systems. Other viruses seem to have followed a similar path. How it interacts with further naive populations would be dispositive, though there may not be any left at this point.

• Updated vaccines are out.
• The race to fusion.
• But solar is the real fusion power.

Aging and Death

Our fate was sealed a very long time ago.

Why do we die? It seems like a cruel and wasteful way to run a biosphere, not to mention a human life. After we have accumulated a lifetime of experience and knowledge, we age, decline, and sign off, whether to go to our just reward, or into oblivion. What is the biological rationale and defense for all this, which the biblical writers assigned to the fairy tale of the snake and the apple?

A recent paper ("A unified framework for evolutionary genetic and physiological theories of aging") discusses evolutionary theories of aging, but in typical French fashion, is both turgid and uninteresting. Aging is widely recognized as the consequence of natural selection, or more precisely, the lack thereof after organisms have finished reproducing. Thus we are at our prime in early adulthood, when we seek mates and raise young. Evolutionarily, it is all downhill from there. In professional sports, athletes are generally over the hill at 30, retiring around 35. Natural selection is increasingly irrelevant after we have done the essential tasks of life- surviving to mate and reproduce. We may participate in our communities, and do useful things, but from an evolutionary perspective, genetic problems at this phase of life have much less impact on reproductive success than those that hit earlier. 

All this is embodied in the "disposable soma" theory of aging, which is that our germ cells are the protected jewels of reproduction, while the rest of our bodies are, well, disposable, and thus experience all the indignities of age once their job of passing on the germ cells is done. The current authors try to push another "developmental" theory of aging, which posits that the tradeoffs between youth and age are not so much the resources or selective constraints focused on germ cell propagation vs the soma, but that developmental pathways are, by selection, optimized for the reproductive phase of life, and thus may be out of tune for later phases. Some pathways are over-functional, some under-functional for the aged body, and that imbalance is sadly uncorrected by evolution. Maybe I am not doing justice to these ideas, which maybe feed into therapeutic options against aging, but I find this distinction uncompelling, and won't discuss it further.

A series of unimpressive distinctions in the academic field studying aging from an evolutionary perspective.

Where did the soma arise? Single cell organisms are naturally unitary- the same cell that survives also mates and is the germ cell for the next generation. There are signs of aging in single cell organisms as well, however. In yeast, "mother" cells have a limited lifespan and ability to put out daughter buds. Even bacteria have "new" and "old" poles, the latter of which accumulate inclusion bodies of proteinaceous junk, which apparently doom the older cell to senescence and death. So all cells are faced with processes that fail over time, and the only sure bet is to start as a "fresh" cell, in some sense. Plants have taken a distinct path from animals, by having bodies and death, yes, but being able to generate germ cells from mature tissues instead of segregating them very early in development into stable and distinct gonads.

Multicellularity began innocently enough. Take slime molds, for example. They live as independent amoebae most of the time, but come together to put out spores, when they have used up the local food. They form a small slug-like body, which then grows a spore-bearing head. Some cells form the spores and get to reproduce, but most don't, being part of the body. The same thing happens with mushrooms, which leave a decaying mushroom body behind after releasing their spores. 

We don't shed alot of tears for the mushrooms of the world, which represent the death-throes of their once-youthful mycelia. But that was the pattern set at the beginning- that bodies are cells differentiated from the germ cells, that provide some useful, competitive function, at the cost of being terminal, and not reproducing. Bodies are forms of both lost energy and material, and lost reproductive potential from all those extra cells. Who could have imagined that they would become so ornate as to totally overwhelm, in mass and complexity, the germ cells that are the point of the whole exercise? Who could have imagined that they would gain feelings, purposes, and memories, and rage against the fate that evolution had in store for them?

On a more mechanistic level, aging appears to arise from many defects. One is the accumulation of mutations, which in soma cells lead to defective proteins being made and defective regulation of cell processes. An extreme form is cancer, as is progeria. Bad proteins and other junk like odd chemicals and chemically modified cell components can accumulate, which is another cause of aging. Cataracts are one example, where the proteins in our lenses wear out from UV exposure. We have quite intricate trash disposal processes, but they can't keep with everything, as we have learned from the advent of modern chemistry and its many toxins. Another cause is more programmatic: senescent cells, which are aged-out and have the virtue that they are blocked from dividing, but have the defect that they put out harmful signals to the immune system that promote inflammation, another general cause of aging.

Aging research has not found a single magic bullet, which makes sense from the evolutionary theory behind it. A few things may be fixable, but mostly the breakdowns were never meant to be remedied or fixed, nor can they be. In fact, our germ cells are not completely immune from aging either, as we learn from older fathers whose children have higher rates of autism. We as somatic bodies are as disposable as any form of packaging, getting those germ cells through a complicated, competitive world, and on to their destination.

• Climate policy as foreign policy.
• The future of home and grid energy.
• Sometimes AI is not so impressive.
• Are Christians better than other Americans?

Oh, to Be Normal

It is a greater accomplishment than commonly appreciated.

The popular media make a fetish of condemning normality. Chase your dream, dare to be different, don't settle for average. Well, that is laudable, and appropriate for the occasional genius, but militates against much larger forces toward uniformity. Just look at styles in clothing, cars, architecture. "Keeping up" with fashions and the times is a marker of, not just normality, but of being alive and part of the larger social community. Achieving normal means not being fossilized in wig and breeches, or bell bottoms. The period of middle school and high school is when these pressures are most acute, as children find places in the wider society, staking their claim with clothing and all the other markers of being "normal". Especially against parents, who have by this time fallen a little back in their ability or desire to keep up with current standards.

But the point I am more interested in is genetic. In genetic terms, normal is typically stated as "wild-type", which is the opposite of mutant. Any particular gene or trait can be construed as normal or defective, with the possibility of being improved in some way over the "wild-type" being exceptionally rare. But summed over an entire genome, one can appreciate that not every gene can be normal. We all have mutations, and thus deviate from normal. In this sense, normality is an impossible, unattainable standard, and as anyone can observe, we all labor under some kind of deficiency. The only question is how severe those deficiencies are, relative to others, and relative to the minimum level of competence we need to survive.

That is where these two threads come together. Young people are continually competing and testing each other for fitness, gauging each other's ability to keep up with the high standard that constitutes "normal" for a culture. It is the beauty queens, and the popular kids, who find themselves at the top of the heap, shining standards of normality in a sea of mediocrity and deficiency. At least until they find out that they might have other, less visible weaknesses, like, perhaps, alcoholism. 

So, not to be all conformist about it, but for all the praise showered on diversity and innovation, there is a lot to be said for standards of normality, which are rather higher than they seem. They actually set significant challenges for everyone to aspire to. They represent, for example, a wide gamut of competencies that undergird society- the ability of people to get along in professional and intimate settings, and the basic knowledge and judgement needed for a democratic political system. Making up for one's deficiencies turns out to be a life-long quest, just as significant as making use of extraordinary gifts or pursuing competitive excellence in some chosen field.

• Ruin here, ruin there.
• The ins and outs of negotiating a large credit bill.
• Notes on early Darwinism.
• "The vast amount of Taiwanese have nothing to fear."

Modeling Cell Division

Is molecular biology ready to use modeling to inform experimental work?

The cell cycle is a holy grail of biology. The first mutants that dissected some of its regulatory apparatus, the CDC mutants of Saccharomyces cerevisiae (yeast), electrified the field and led to a Nobel prize. These were temperature sensitive mutants, making only small changes to the protein sequence that rendered that protein inactive at high temperature (thus inducing a cell cycle arrest phenotype), while allowing wild-type growth at normal temperatures. In the fifty years since, a great deal of the circuitry has been worked out, with the result that it is now possible, as a recent paper describes, to make a detailed mathematical model of the process that claims to be useful in the sense of explaining existing findings in a unified model and making predictions of places to look for additional actors.

At the center of this regulatory scheme are transcription activators, SBF/MBF, that are partly controlled by, and in turn control the synthesis of, a series of cyclins. Cyclins are proteins that were observed (another Nobel prize) to have striking variations in abundance during the cell cycle. There are characteristic cyclins for each phase of the cell cycle, which goes from G1, a resting phase, to S, which is DNA replication, to G2, a second resting phase, and then M, which is mitosis, which brings us back to G1. Cyclins work by binding to a central protein kinase, Cdc28, which, as regulated by each distinct cyclin, phosphorylates and thus regulates distinct sets of target proteins. The key decision a cell has to make is whether to commit to DNA replication, i.e. S phase. No cell wants to run out of energy during this process, so its size and metabolic state needs to be carefully monitored. That is done by Cyclin 3 (Cln3), Whi5, and Bck2, which each influence whether the SBF/MBF regulators are active. 

Some highly simplified elements of the yeast cell cycle. Cyclins (Cln and Clb) are regulators of a central protein kinase, Cdc28, that direct it to regulate appropriate targets at each stage of the cell cycle. Cyclins themselves are regulated by transcriptional control (here, the activators SBF and MBF), and then destroyed at appropriate times by proteolysis, rendering them abundant only at specific times during the cell cycle. Focusing on the "START" process that starts the process from rest (G1 phase) to new bud formation and DNA replication (S phase), Cln3 and Bck2 respond to upstream nutritional and size cues, and each activate the SBF/MBF transcription activator.

As outlined in the figure above, Cyclin 3 is the G1 cyclin, which, in complex with Cdc28 phosphorylates Whi5, turning it off. Whi5 is an inhibitor that binds to SBF/MBF, so the Cyclin 3 activation turns these regulators on, and thus starts off the cell cycle under the proper conditions. Incidentally, the mammalian version of Whi5, Rb (for retinoblastoma), is a notorious oncogene, that, when mutated, releases cells from regulatory control over cell division. SBF and MBF bind to genes for the next series of cyclins, Cln1, Cln2, Clb5, Clb6. The first two are further G1 cyclins that orchestrate the end of G1. They induce phosphorylation and inactivation of Sic1 and Cdc6, which are inhibitors of Clb5 and Clb6. These latter two are then the initiators of S phase and DNA replication. Meanwhile, Cln3 stays around till M phase, but is then degraded in definitive fashion by the proteases that end M phase. Starvation conditions lead to rapid degradation of Cln3 at all times, and thus to no chance of starting a new cell cycle.

Charts of the abundance of some cyclins through the cell cycle. Each one has its time to shine, after which it is ubiquitinated and sent off to the recycling center / proteasome.

Bck2 is another activator of SBF/MBF that is unrelated to the Cln3/Whi5 system, but also integrates cell size and metabolic status information. Null mutants of Cln3 (or Bck2) are viable, if altered in cell cycle, while double null mutants of Cln3 and Bck2 are dead, indicating that these regulators are each important, in a complementary way, in cell cycle control. Given that little is known about Bck2, the modelers in this paper assume various properties and hope for the best down the line, predicting that cell size (at the key transition to S phase) is more affected in the Cln3 null mutant than in the Bck2 null mutant, since in the former, excess active Whi5 soaks up most of the available SBF/MBF, and requiring extra-high and active levels of Bck2 to overcome this barrier and activate the G1 cyclins and other genes.

The modelers are working from the accumulated, mostly genetic data, and in turn validate their models against the same genetic data, plus a few extra mutants they or others have made. The models are mathematical representations of how each node (i.e protein, or gene) in the system responds to the others, but since there are a multitude of unknowns, (such as what really regulates Bck2 from upstream, to cite just one example), the system is not really able to make predictions, but rather fine-tunes/reconciles what knowledge there is, and, at best, points to gaps in knowledge. It is a bit like AI, which magically recombines and regurgitates material from a vast corpus based on piece-wise cues, but is not going to find new data, other than through its notorious hallucinations.

For example, a new paper came out after this modeling, which finds that Cln3 affects Cln2 abundance by mechanisms quite apart from its SBF/MBF transcriptional control, and that it regulates cell size in large part at M phase, not through its G1/S gating. All this comes from new experimental work, unanticipated by the modeling. So, in the end, experimental work always trumps modeling, which is a bit different than how things are in, say, physics, where sometimes the modeling can be so strong that it predicts new particles, forces, and other phenomena, to be validated later experimentally. Biology may have its master predictive model in the theory of evolution, but genetics and molecular biology remain much more of an empirical slog through the resulting glorious mess.

• Bitcoin isn't a currency, but rather just another asset class, one without any fundamental or socially positive value. A little like gold, actually, except without gold's resilience against social / technological disruption.
• The disastrous post-Soviet economic transition, on our advice.
• The enormous labor drain, and resource drain, from global South to North.

Welcome to Lubyanka!

Another case of penal systems illuminating their culture.

Most of Aleksandr Solzhenitsyn's In the First Circle is a desultory slog, at least if you have already read the Gulag Archipelago. But there are a few glorious set-pieces. One is the mock trial of Prince Igor of Kiev that the prisoners stage in their free time, a bitter satire of the Soviet judicial system. The second is a meticulous description of how prisoners are brought into and introduced to the Lubyanka prison- the central prison of the KGB/FSB/Cheka/GPU/OGPU/NKVD/NKGB, etc.. the frequently renamed, but never-changing organ of the Russian government.

The character is Innokenty Volodin, a Soviet diplomat who has recently had second thoughts about the rightness of the Soviet system, and has placed a call (around which the book's plot, such as it is, mostly revolves) to the Americans to prevent Russia from obtaining certain critical atomic secrets. Solzhenitsyn carefully prepares the way by portraying Volodin's rarified position and luxurious life. As was customary, Volodin is lured into his arrest under false pretenses, and finds himself driven to the prison almost before he knows what has happened. Then, with almost loving detail, Solzhenitsyn describes the not just systematic, but virtuosic process of degradation, step by step, shred by shred, of Volodin's humanity, as he is inducted into Lubyanka.

One cardinal rule is that prisoners must have no contact with other prisoners. Even to see others is forbidden. As they are conducted from one cell to the next, they are shoved into mini phone-booth cells if another prisoner is being conducted in the opposite direction. Their possessions are gradually taken away, down to buttons, belts, and steel shoe shanks. They are shorn. They are sleep deprived. They are relentlessly illuminated by glaring bulbs. They are spied on constantly. They are moved relentlessly from place to place and disoriented. In the middle of the night, the building is abuzz with activity, as though this were the very nerve center of the Soviet empire. 

While the rest of Russian society is mired, or cowed, in mediocrity, this is a shining point of competence. The purest expression of its obsessive leader, and the product of decades of careful study and accumulated wisdom. It is also a deeper expression of the nature of Russian society- its reflexive despotism and its strange infatuation with suffering. The closest thing we have is mafia culture, with its honor codes, brutality, and constant battle for dominance. Chess, the emblematic game of Russia, expresses this view of life as a pitiless contest to crush one's opponent. There may be a lot of historical reasons for this nature, such as the long centuries of Mongol rule, the many invasions, both ancient and modern, and the perceived success of leaders such as Ivan the Terrible and Stalin, but it is a deep and disturbing aspect of the Russian psyche. 

Should we have expected anything else, in the long road of declining relations after the cold war? Should the Russian people give thanks to the ruthlessness of their national leadership and psyche for the current position of relative power they wield in the world, far out of proportion to their population or economic strength? Other countries with larger populations peacefully mind their own business, avoid outside entanglements, and eschew invading their neighbors. It is the bullies, the intransigent, and the cruel, who appear to account for most of the drama in the world. Should we understand them, or fight against them?

• Another way to crush spirits.
• It is good to be outside.
• AI is not helping job seekers.
• On paying the devil his due.
• Why have kids anymore?

Putting Body Parts in Their Places

How HOX genes run development, on butterfly wings.

I have written about the HOX complex of genes several times, because they constitute a grail of developmental genetics- genes that specify the identity of body parts. They occupy the middle of a body plan cascade of gene regulation, downstream from broader specifiers for anterior/posterior orientation, regional and segment specification, and in turn upstream of many more genes that specify the details of organ and tissue construction. Each of the HOX genes encodes a transcriptional regulator, and the name of one says it all- antennapedia. In fruit flies, where all this was first discovered, loss of antennapedia converts some legs into antennae, and extra expression of antennapedia converts antennae on the head into legs.

The HOX complex (named for the homeobox DNA binding motif of the proteins they encode) is linear, arranged from head-affecting genes (labial, proboscipedia) to abdomen-affecting genes (abdominal A, abdominal B; evidently the geneticist's flair for naming ran out by this point). This arrangement is almost universally conserved, and turns out to reflect molecular mechanisms operating on the complex. That is, it "opens" in a progressive manner during development, on the chromosome. Repression of chromatin is a very common and sturdy way to turn genes off, and tends to affect nearby genes, in a spreading effect. So it turns out to be easy, in some sense, to set up the HOX complex to have this chromatin repression lifted in a segmental fashion, by upstream regulators, whereby only the head sections are allowed to be expressed in head tissues, but all the genes are allowed to be expressed in the final abdominal segment. That is why the unexpected expression of antennapedia, which is the fifth of eight HOX genes, in the head, leads to a thoracic tissue (legs) forming on the head.

A recent paper delved a little more deeply into this story, using butterflies, which have a normal linearly conserved HOX cluster and are easy to diagnose for certain body part transformations (called homeotic) on their beautiful wings. The main thing these researchers were interested in is the genetic elements that separate one part of the HOX cluster from other parts. These are boundary or "insulator" elements that separate topologically associated domains (called TADs). Each HOX gene is surrounded by various regulatory enhancer and inhibitor sites in the DNA that are bound by regulatory proteins. And it is imperative that these sites be directed only to the intended gene, not neighboring genes. That is why such TADs exist, to isolate the regulation of genes from others nearby. There are now a variety of methods to map such TADs, by looking where chromatin (histones) are open or closed, or where DNA can be cut by enzymes in the native chromatin, or where crosslinks can be formed between DNA molecules, and others.

The question posed here was whether a boundary element, if deleted, would cause a homeotic transformation in the butterflies they were studying. They found, unfortunately, that it was impossible to generate whole animals with the deletions and other mutations they were engineering, so they settled for injecting the CRISPER mutational molecules into larval tissues and watching how they affected the adults in mosaic form, with some mutant tissues, some wild-type. The boundary they focused on was between antennapedia (Antp) and ultrabithorax (Ubx), and the tissues the forewings, where Ubx is normally off, and hindwings, where Ubx is normally on. Using methods to look at the open state of chromatin, they found that the Ubx gene is dramatically opened in hindwings, relative to forewings. Nevertheless, the boundary remains in place throughout, showing that there is a pretty strong isolation from Antp to Ubx, though they are next door and a couple hundred thousand basepairs apart. Which in genomic terms is not terribly far, while it leaves plenty of space for enhancers, promotes, introns, boundary elements, and other regulatory paraphernalia.

Analysis of the site-to-site chromosomal closeness and accessibility across the HOX locus of the butterfly Junonia coenia. The genetic loci are noted at the bottom, and the site-to-site hit rates are noted in the top panels, with blue for low rates of contact, and orange/red for high rates of contact. At top is the forewing, and at bottom is the hindwing, where Ubx is expressed, thus the high open-ness and intra-site contact within its topological domain (TAD). Yet the boundary between Ubx and Anp to its left (dotted lines at bottom) remains very strong in both tissues. In green is a measure of transcription from this DNA, in differential terms hindwing minus forewing, showing the strong repression of Ubx in the forewing, top panel.

The researchers naturally wanted to mutate the boundary element, (Antp-Ubx_BE), which they deduced lay at a set of binding sites (featuring CCCTC) for the protein CTCF, a well-known insulating boundary regulator. Note, interestingly, that in the image above, the last exon (blue) of Ubx (transcription goes right to left) lies across the boundary element, and in the topological domain of the Antp gene. This means that while all the regulatory apparatus of Ubx is located in its own domain, on the right side, it is OK for transcription to leak across- that has no regulatory implications. 

Effects of removing the boundary element between Ubx and Antp. Detailed description is in the text below. 

Removal of this boundary element, using CRISPER technology in portions of the larval tissues, had the expected partial effects on the larval, and later adult, wings of this butterfly. First, note that in panel D insets, the wild type larval forewing shows no expression of Ubx, (green), while the wild type hind wing shows wide-spread expression. This is the core role of the HOX locus and the Ubx gene- locate its expression in the correct body parts to then induce the correct tissues to develop. The larval wing tissue of the mosaic mutant, also in D, shows, in the forewing, extensive patchy expression of Ubx. This is then reflected in the adult (different animals) in the upper panels, in the mangled eyespot of the fully formed wing (center panel, compared to wild-type forewing and hindwing to each side). It is a small effect, but then these are small mutations, done in only a fraction of the larval cells, as well.

So here we are, getting into the nuts and bolts of how body parts are positioned and encoded. There are large regions around these genes devoted to regulatory affairs, including the management of chromatin repression, the insulation of one region from another, the enhancer and repressor sites that integrate myriad upstream signals (i.e. other DNA binding proteins) to come up with the detailed pattern of expression of these HOX genes. Which in turn control hundreds of other genes to execute the genetic program. This program can hardly be thought of as a blueprint, nor a "design" in anyone's eye, divine or otherwise. It resembles much more a vast pile of computer code that has accreted over time with occasional additions of subroutines, hacks, duplicated bits, and accidental losses, adding up to a method for making a body that is robust in some respects to the slings and arrows of fortune, but naturally not to mutations in its own code.

• How it looks from abroad.
• Was Mawmaw paranoid, or bipolar?
• China is plowing its investment excess into renewable energy.
• Visions of man.
• The health insurance system is dysfunctional.

Hungary for Power

Hungary has become a one-party, authoritarian state, not a democracy.

Victor Orban recently paid a visit to Donald Trump in Florida, with glowing photos and pledges of goodwill. Republicans in the US have nurtured a deep fascination and alliance with Orban and his government, holding several CPAC conventions in Hungary, and hosting Orban and his lieutenants at US events. It is clear that they view Hungary as a shining example and template of where they could take the US. Not the shining city on a hill of Reagan's democratic and anti-authoritarian dreams, but a whole other kind of city, one that never will fall into Democratic hands again.

So it is worth looking in detail at what has happened in Hungary, to observe the ideals of our current Republicans and what is in store for the rest of us from a second Trump term. I was, incidentally, beaten to the punch of this analysis by a recent story in the Atlantic. Orban's party, Fidesz, is very similar to the GOP in its mix of business right-wingery and rural values. Its strength is handing out the red meat of traditional, anti-cosmopolitan values to the rural base, along with helpful economic subsidies. In the pivotal 2018 election, it won all the rural districts, even though the opposition bowed to the logic of re-written (winner-take-all) electoral system and tried to join into a unified party. 

Fidesz came to power originally on an anti-socialist platform, vowing to get rid of the remnant bits of the prior communist system, which had settled into the same kind of semi-kleptocratic mode as in most of the former Soviet states and its satellites. That they did, but not to bring an end to corruption, let alone authoritarianism, but rather to partake themselves instead. After coming into power, Fidesz rewrote the constitution, in ways large and small to entrench their own power, and has since continued a campaign of extremely effective, gradual, and often surreptitious legislation to cement its advantages. Gerrymandering is now standard procedure, which when combined with the winner-take-all districts creates the opportunity to win overwhelming majorities in parliament founded on very thin electoral pluralities. Small parties can not win any more, but are also prohibited from combining with other small parties into election list coalitions.

The courts were remade by putting them under the control of a political appointee- the president of the National Judicial Office. This president is appointed by parliament, and in turn appoints, promotes, and runs the operations and budget of the whole judicial system. Needless to say, it is heavily influenced by the now Fidesz-controlled parliament and executive.

The media has been remade by gradual pressure on independent media owners to sell to Fidesz-friendly interests, which now control 90% of the country's media. Government advertising buys were strategically placed with friendly outlets, and government run media was put under direct political control. A Russian inspired "security" law was passed to outlaw ill-defined criticism of the state, public morality, or "imbalance" of coverage, answerable naturally to a parliamentary-appointed body, rather than the courts. Imagine if in the second Trump administration, PBS and NPR were put under political control and given a "FOX" makeover. 

Hungary is now effectively a one-party authoritarian state with managed elections. We are not far off. To see the battle of titanic interests and billionaires now openly showering money on favored candidates, and extending their tentacles down to the school board level, is sickening. The Republican party has partnered with Heritage foundation to offer an openly Orbanist plan for the second Trump administration. The court system has already re-written our constitution in extensive ways over the last four years, without an amendment being passed, or even proposed. The antics of Judge Eileen Cannon show that very little may remain of the rule of law if it is left in the hands of partisan extremists.

And our media is in even more perilous condition, with the relentless lying of FOX, Sinclair, and their ecosystem. The Republican convention just past was a pageant of lies and grift, betokening the criminal enterprise that party has turned into. Headed by their adored, and now divine, leader who is not just a felon and business fraud, but rapist and insurrectionist as well. But no matter. With enough money, and enough shamelessness, anything is possible.

• Everything is fine!
• Scandals erupt out of a corrupted system.
• The new GOP.
• Which is a blue collar party, right?
• Tom Tomorrow, on last week.
• Historical note on the great California land grab, and the fate of "free" land generally.
• Notes from the early days of life.

The Long Tail of Genome Duplication

A new genomic sequence of hagfish tells us a little about our origins.

Hagfish- not a fish, and not very pretty, but it occupies a special place in evolution, as a vertebrate that diverged very early (along with lampreys, forming the cyclostome branch) from the rest of the jawed vertebrates (the gnathostome branch). The lamprey has been central to studies of the blood clotting system, which is a classic story of gradual elaboration over time, with more steps added to the cascade, enabling faster clotting and finer regulation.

A highly schematic portrayal (not to scale!) of the evolutionary history of animal life on earth.

A recent paper reported a full genome sequence of hagfish, and came up with some interesting observations about the history of vertebrate genomes. At about three billion nucleotides, this genome is about as large as ours. (Yet again, size doesn't see, to matter much, when it comes to genomes.) They confirm that lampreys and hagfish make up a single lineage, separate from all other animals and especially from the jawed vertebrates. For example, though lampreys have 84 chromosomes to the hagfish's 17, this resulted from repeated splitting of chromosomes, and each lamprey chromosome can be mostly mapped to one hagfish chromosome, accepting that a lot of other gene movement and change has taken place in the roughly 460 million years since these lineages diverged. 

Hagfish (bottom) and lamprey (top) chromosomes pretty much line up, indicating that despite the splitting of the lamprey genome, there hasn't been a great deal of shuffling over the intervening 460 million years.

The most important parts of this paper are on the history of genome duplications that happened during this early phase of vertebrate evolution. Whole genome duplications are an extremely powerful engine of change, supplying the organism with huge amounts of new genetic material. Over time, most of the duplicated genes are discarded again (in a process they call re-diploidization). But many are not, if they have gained some foothold in providing more of an important product, or differentiated themselves from each other in some other way. Our genomes are full of families, some extremely large, of related genes that have finely differentiated functions. Many of these copies originated in long-ago genome duplications, while others originated in smaller duplication accidents. It is startling to hear from self-labeled scientists in the so-called intelligent design movement that there is some rule or law against such copying of information, by their ridiculous theories of specified information. Hagfish certainly never heard of such a thing.

At any rate, these researchers confirm that the earliest vertebrate lineage, around 530 million years ago, experienced two genome duplications which led to a large increment of new genes and evolutionary innovation. What they find now is that the cyclostome lineage experienced another genome three-fold duplication (near its origin, about 460 million years ago, leading to another round of copies and innovation. And lastly, the gnathostome lineage separately experienced its own genome four-fold duplication around the same time, after it had diverged from the cyclostome lineage. One might say that the gnathostomes made better use of their genomic manna, generating jaws, teeth, ears, thymus, better immune systems, and the other features that led them to win the race of the animal kingdom. But hagfish are still around, showing that primitive forms can find a place in the scheme of things, as the biosphere gets larger and more diverse over time.

A classic example of gene replication is the Hox cluster, which are a set of genes that have the power of dictating what body part occurs where. They are gene regulators that function in the middle of the developmental sequence, after determination of the overall body axis and segmentation, and themselves regulating downstream genes governing features as they occur in different segments, such as limbs, parts of the head, fingers, etc. Flies have one Hox cluster, split into two parts. The extremely primitive chordate amphioxus, which far predates the cyclostomes, also has one complete Hox cluster, as diagrammed below. Most other vertebrates, including us, have four Hox clusters, amounting to over thirty of these transcription regulators. These four clusters arose from the inferred genome duplications very early in the vertebrate lineage, prior to the advent of the cyclostomes. 

Hox clusters and their origins, as inferred by the current authors. The red/blue points at the left mark whole genome duplications (or more) that have been inferred by these or other authors. More description is in the main text below.

The inferred genome duplications during early chordate evolution, noted on the far left of the diagram above, led to duplicated clusters of Hox genes. Amphioxus (top) is the earliest branching chordate, and has only one full Hox cluster of transcription regulators, which, in general terms, control, during development, the expression of body parts along the body axis, with the order of genes in the cluster paralleling expression and action along the body axis. Chicken as a gnathostome has four copies of the cluster, with a few of the component genes lost over time. Hagfish have six copies of this Hox cluster, some rather skeletal, stemming from its genome duplication events. Clearly several whole clusters have also been lost, as in some cases the genome duplications experienced by the cyclostomes resolved back to diploidy without leaving an extra copy of this cluster. The net effect is to allow all these organisms greater options for controlling the identity and form of different parts of the body, particularly, in the case of gnathostomes, the head.

Genome duplications are one of those fast events in evolution that are highly influential, unlike the usual slow and steady selection and optimization that is the rule in the Darwinian theory. Unlike mass extinction, another kind of fast event in evolution, genome duplications are highly constructive, providing fodder on a mass (if microscopic) scale for new functions and specializations that help account for some of the more rapid events in the history of life, such as the rise of chordates and then vertebrates in the wake of the Cambrian explosion.

• America doesn't work for workers.
• Diagnosis Biden.
• Don't use rat poison.