Dangerous Memories

Some memory formation involves extracellular structures, DNA damage, and immune component activation / inflammation.

The physical nature of memories in the brain is under intensive scrutiny. The leading general theory is that of positive reinforcement, where neurons that are co-activated strengthen their connections, enhancing their ability to co-fire and thus to express the same pattern again in the future. The nature of these connections has been somewhat nebulous, assumed to just be the size and stability of their synaptic touch-points. But it turns out that there is a great deal more going on.

A recent paper started with a fishing expedition, looking at changes in gene expression in neurons at various time points after the mice were subjected to a fear learning regimen. They took this out to much longer time points (up to a month) than had been contemplated previously. At short times, a bunch of well-known signals and growth-oriented gene expression happened. At the longest time points, organization of a structure called the perineural net (PNN) was read out of the gene expression signals. This is a extracellular matrix sheath that appears to stabilize neuronal connections and play a role in long-term memory and learning. 

But the real shocker came at the intermediate time point of about four days. Here, there was overexpression of TLR9, which is an immune system detector of broken / bacterial DNA, and inducer in turn of inflammatory responses. This led the authors down a long rabbit hole of investigating what kind of DNA fragmentation is activating this signal, how common this is, how influential it is for learning, and what the downstream pathways are. Apparently, neuronal excitation, particularly over-excitation that might be experienced under intense fear conditions, isn't just stressful in a semiotic sense, but is highly stressful to the participating neurons. There are signs of mitochondrial over-activity and oxidative stress, which lead to DNA breakage in the nucleus, and even nuclear perforation. It is a shocking situation for cells that need to survive for the lifetime of the animal. Granted, these are not germ cells that prioritize genomic stability above all else, but getting your DNA broken just for the purpose of signaling a stress response that feeds into memory formation? That is weird.

Some neuronal cell bodies after fear learning. The red dye is against a marker of DNA repair proteins, which form tight dots around broken DNA. The blue is a general DNA stain, and the green is against a component of the nuclear envelope, showing here that nuclear envelopes have broken in many of these cells.

The researchers found that there are classic signs of DNA breakage, which are what is turning on the TLR9 protein, such as seeing concentrated double-strand DNA repair complexes. All this stress also turned on proteases called caspases, though not the cell suicide program that these caspases typically initiate. Many of the DNA break and repair complexes were, thanks to nuclear perforation, located diffusely at the centrosome, not in the nucleus. TLR9 turns on an inflammatory response via NFKB / RELA. This is clearly a huge event for these cells, not sending them into suicide, but all the alarms short of that are going off.

The interesting part was when the researchers asked whether, by deleting the TLR9 or related genes in the pathway, they could affect learning. Yes, indeed- the fear memory was dependent on the expression of this gene in neurons, and on this cell stress pathway, which appears to be the precondition of setting up the perineural net structures and overall stabilization. Additionally, the DNA damage still happened, but was not properly recognized and repaired in the absence of TLR9, creating an even more dangerous situation for the affected neurons- of genomic instability amidst unrepaired DNA.

When TRL9 is knocked out, DNA repair is cancelled. At bottom are wild-type cells, and at top are mouse neurons after fear learning that have had the gene TLR9 deleted. The red dye is against DNA repair proteins, as is the blue dye in the right-most frames. The top row is devoid of these repair activities.

This paper and its antecedent literature are making the case that memory formation (at least under these somewhat traumatic conditions- whether this is true for all kinds of memory formation remains to be seen) has commandeered ancient, diverse, and quite dangerous forms of cell stress response. It is no picnic in the park with madeleines. It is an all-hands-on-deck disaster scene that puts the cell into a permanently altered trajectory, and carries a variety of long-term risks, such as cancer formation from all the DNA breakage and end-joining repair, which is not very accurate. They mention in passing that some drugs have been recently developed against TLR9, which are being used to dampen inflammatory activities in the brain. But this new work indicates that such drugs are likely double-edged swords, that could impair both learning and the long-term health of treated neurons and brains.

• Cutting the line to the American Dream.

Cooperation is Very Hard

Few animals engage in productive cooperation outside family kin groups.

It might be hard to imagine in our current political climate, but cooperation is the core trait of modern humans- the ability to form groups, make rules, exchange favors, and get big things done. We even cooperate for fleeting efforts with people we will never meet again. But theories explaining cooperation have been hard-won and so far quite limited in evolutionary biology. Kin selection is perhaps the only serious theory of this type, making cooperation a strict function of shared genes, which in turn sees its role rapidly diminishing in larger groups of organisms, with the exception of peculiar families like the social insects. 

Two other explanations for cooperative behavior have been developed- repeated interaction, and group selection. In the first, assuming that humans evolved in small groups, everyone knew everything about everyone, so reputation is everything, and thus cooperation within a group is the default state. In the second, different groups with different levels of in-group cooperation would fight it out in some form, perhaps militarily, leading to the success of better-cooperating groups. A recent paper (with review) used improved modeling to suggest that neither of these two explanations holds much water on its own, but in particular ways could be combined into something they call "super-additive cooperation". I.e. human society.

The key modeling advance here was to use graded rather than binary functions for interaction rewards. Likewise, they also allowed other forms of cooperation to compete with reciprocal cooperation. This allowed subjects and modeled entities to do what people always do- get away with giving a little less in return, which sends the whole game sliding into oblivion. That is, unless you are known to others in your own group, in which case, getting and giving positive rewards becomes a virtuous cycle with ever-increasing payoffs, thus the term super-additive. The combination of in-group membership and repeated interactions provides the magic. 

Detailed modeling of cooperation (termed "escalation" of cooperation) under some key conditions. Top is repeated interactions without group selection. Next is group competition without repeated in-group interactions. Third is the joint combination. The legend at top right ranges from generous cooperation at top to selfishness at the bottom. This is modeled as money transfers between participants, which are tallied in the leftmost graphs, and fall to minimal levels over time in the top two scenarios.

But does this amount to group selection? These authors suggest that, as typically understood, group selection is not very strong and not strong enough to support the evolution of cooperation. Among humans, conflicting groups are genetically different to only infinitesimal degrees. Migration and intermarriage (forced and otherwise) are so frequent that it would be practically impossible to build selectable differences over the needed time scales. On the other hand, human societies exhibit cultural variation as well, and this kind of variation is more extensive and much more rapidly developing than genetic variation, creating differences between groups that can withstand moderate levels of migration and remain distinctive and selective. As cultural group selection, this is not the same as group selection in classic evolutionary theory, and indeed, it may be hard to relate this to evolutionary theory at all. But it certainly leads to differential survival and reproduction, whatever the genetic background to the cooperative, group feeling, and other traits that feed into the culture.

"We also show that combining the two mechanisms generates strong positive interactions. Positive interactions occur because intergroup competitions can stabilize ingroup cooperation against ambiguous reciprocity, and intergroup competitions often do this even when they do not support cooperation on their own. When the mechanisms interact, the result is the evolution of cooperative reciprocity with ingroup members, which amplifies cooperation within groups, and uncooperative reciprocity with outgroup members, which erodes cooperation between groups."

...

"Group competition can change the balance of forces by adding a mechanism that favours relatively cooperative groups. The higher payoffs associated with escalation can now dominate the fragility of escalation, with the final outcome a cooperative escalating equilibrium. When group competition shifts the balance in this way, the cooperative outcome does not require large differences between groups."

Humans and their culture are extremely complex, and this is hardly the last word on mechanisms of cooperation, which include surveillance, punishment, and much else. But at least this study can dispose of the simpler evolutionary explanations, that are accessible to uncultured organisms and explain why free cooperation among unrelated individuals is limited out in nature, to behaviors with immediate paybacks like schooling, herding, flocking, and nutrient exchanges.

• Market-origin theory for Covid gets more support.

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.