At least sometimes.. how replication of the centrosome creates asymmetry of cell division as a whole, and what makes that asymmetry happen.
Cell fates can hinge on very simple distinctions. The orientation of dividing cells in a tight compartment may force one out of the cozy home, and into a different environment, which induces differentiation. Stem cells, those notorious objects of awe and research, are progenitor cells that stay undifferentiated themselves, but divide to produce progeny that differentiate into one, or many, different cell types. At the root of this capacity of stem cells is some kind of asymmetric cell division, whether enforced physically by an environmental micro-niche, or internally by molecular means. And a dominant way for cells to have intrinsic asymmetry is for their spindle apparatus to lead the way. Our previous overview of the centrosome (or spindle pole body) described its physical structure and ability to organize the microtubules of the cell, particularly during cell division. A recent paper discussed how the centrosome itself divides and originates a basic asymmetry of all eukaryotic cells.
The centrosome is a complicated structure that replicates in tandem with the rest of the cell cycle. Centrosomes do not divide in the middle or by fission. Rather, the daughter develops off to the side of the mother. Centrosomes are embedded in the nuclear envelope, and the mother develops a short extension, called a bridge or half-bridge, off its side, off of which the daughter develops, also anchored in the nuclear envelope. Though there are hundreds of associated proteins, the key components in this story are NUD1, which forms part of the core of the centrosome, and SPC72, which binds to NUD1 and also binds to the microtubules (made of the protein tubulin) which it is the job of the centrosome to organize. In yeast cells, which divide into very distinct mother and daughter (bud) cells, the mother centrosome (called the spindle pole body) leads the way into division and always goes into the daughter cell, while the daughter centrosome stays in the mother cell.
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| The deduced structure of some members of the centrosome/spindle pole in yeast cells. Everything below the nuclear envelope is inside the nucleus, while everything above is in the cytoplasm. The proteins most significant in this study are gamma tubulin (yTC), Spc72, and Nud1. OP stands for outer plaque, CP central plaque, IP inner plaque, as these structures look like separate dense layers in electron microscopy. To the right side of the central plaque is a dark bit called the half-bridge, on the other side of which the daughter centrosome develops, during cell division. |
The authors asked why this difference exists- why do mother centrosomes act first to go to the outside of the cell where the bud forms? Is it simply a matter of immaturity, that the daughter centrosome is not complete at this point, (and if so, why), or is there more specific regulation involved that enforces this behavior? They use a combined approach in yeast cells combining advanced fluorescence microscopy with genetics to find the connection between the cell cycle and the progressive development of the daughter centrosome.
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| Yeast cells with three mutant centrosome proteins, each engineered as fusions to fluorescent proteins of different color, were used to show the relative positions of KAR1, (green), which lies in the half-bridge between the mother and daughter centrosomes. Three successive cell cycle states are shown. Spc42, (blue), at the core of the centrosome, and gamma tubulin (red; Tub4, or alternately Spc72, which lies just inside Tup4), which is at the outside and mediates between the centrosome and the tubulin-containing microtubules. Note that the addition of gamma tubulin is a late event, after Spc42 appears in the daughter. The bottom series is oriented essentially upside down vs the top two series. |
What they find, looking at cells going through all stages of cell division, is that the assembly of the daughter centrosome is stepwise, with inner components added before outer ones. Particularly, the final structural elements of Spc72 and gamma tubulin wait till the start of anaphase, when the cells are just about to divide, to be added to the daughter centrosome. The authors then bring in key cell cycle mutants to show that the central controller of the cell cycle, cyclin-dependent kinase CDK, is what is causing the hold-up. This kinase (a protein that phosphorylates other proteins, as a means of regulation) orchestrates much of the yeast cell cycle, as it does in all eukaryotic cells, subject to a blizzard of other regulatory influences. They observed that special inducible mutations (sensitive versions of the protein that shut off at elevated temperature) of CDK would stop this spindle assembly process, suggesting that some component was being phosphorylated by CDK at the key time of the cell cycle. Then, after systematically mutating possible CDK target phosphorylation sites on likely proteins of the centrosome, they came up with Nud1 as the probable target of CDK control. This makes complete sense, since Spc72 assembles on top of Nud1 in the structure, as diagrammed at top. They go on to show the direct phosphorylation of Nud1 by CDK, as well as direct binding between Nud1 and Spc72.
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| Final model from the article shows how the mechanics they revealed relate to the cell cycle. A daughter centrosome slowly develops off the side of the mother centrosome, but its "licensing" by CDK to nucleate microtubules (black rods anchored by the blue cones) only comes later on in M phase, just as the final steps of division need to take place. This gives the mother centrosome the jump, allowing it to migrate to the bud (daughter cell) and nucleate the microtubules needed to drive half of the replicated DNA/chromosomes into the bud. GammaTC is nucleating gamma tubulin, "P" stands for activating phosphorylation sites on Nud1. |
This is a nice example of the power of a model system like yeast, whose rich set of mutants, ease of genetic and physical manipulation, complete genome sequence and associated bioinformatics, and many other technologies make it a gold mine of basic research. The only hard part was the microscopy, since yeast cells are substantially smaller than human cells, making that part of the study a tour de force.
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