Decreasing oocyte competence with maternal aging is a major factor in human infertility. To investigate the age-dependent molecular changes in a mouse model, we compared the expression profiles of metaphase II oocytes collected from 5- to 6-week-old mice with those collected from 42- to 45-week-old mice using the NIA 22K 60-mer oligo microarray. Among approximately 11,000 genes whose transcripts were detected in oocytes, about 5% (530) showed statistically significant expression changes, excluding the possibility of global decline in transcript abundance. Consistent with the generally accepted view of aging, the differentially expressed genes included ones involved in mitochondrial function and oxidative stress. However, the expression of other genes involved in chromatin structure, DNA methylation, genome stability and RNA helicases was also altered, suggesting the existence of additional mechanisms for aging. Among the transcripts decreased with aging, we identified and characterized a group of new oocyte-specific genes, members of the human NACHT, leucine-rich repeat and PYD-containing (NALP) gene family. These results have implications for aging research as well as for clinical ooplasmic donation to rejuvenate aging oocytes.

We developed a stochastic model of microtubule (MT) assembly dynamics that estimates tubulin-tubulin bond energies, mechanical energy stored in the lattice dimers, and the size of the tubulin-GTP cap at MT tips. First, a simple assembly͞disassembly state model was used to screen possible combinations of lateral bond energy (⌬G Lat) and longitudinal bond energy (⌬GLong) plus the free energy of immobilizing a dimer in the MT lattice (⌬G S) for rates of MT growth and shortening measured experimentally. This analysis predicts ⌬G Lat in the range of ؊3.2 to ؊5.7 kBT and ⌬GLong plus ⌬GS in the range of ؊6.8 to ؊9.4 kBT. Based on these estimates, the energy of conformational stress for a single tubulin-GDP dimer in the lattice is 2.1-2.5 k BT. Second, we studied how tubulin-GTP cap size fluctuates with different hydrolysis rules and show that a mechanism of directly coupling subunit addition to hydrolysis fails to support MT growth, whereas a finite hydrolysis rate allows growth. By adding rules to mimic the mechanical constraints present at the MT tip, the model generates tubulin-GTP caps similar in size to experimental estimates. Finally, by combining assembly͞ disassembly and cap dynamics, we generate MT dynamic instability with rates and transition frequencies similar to those measured experimentally. Our model serves as a platform to examine GTPcap dynamics and allows predictions of how MT-associated proteins and other effectors alter the energetics of MT assembly.M icrotubule (MT) dynamic instability plays a critical role in chromosome movement and separation during mitosis. MTs grow, shorten, and transition between these states at rates governed by the presence of various MT effectors, such as divalent cations, MT-associated proteins (MAPs), and drugs such as Taxol (1).MTs are composed of heterodimer subunits of ␣-and ␤-tubulin. A guanine nucleotide (GTP or GDP) is positioned on the ␤ monomer opposite the interface between the two monomers, where it is hydrolyzable (if it is GTP) and exchangeable. The ␤ monomer end of the dimer faces the (ϩ) end of the MT, which is the end of more active dynamics and kinetochore attachment during cell division. The ␣ monomer faces the (Ϫ) end of the MT, which originates at the centrosome. MTs are thought to transition from a state of growth to a state of rapid shortening, termed a ''catastrophe'', when the tubulin-GTP ''cap'' is stochastically lost from the tip of the MT. The tubulin-GTP cap must exist because it has been demonstrated that tubulin-GTP subunits are added to the ends during assembly. A tubulin-GTP cap, however, has not been detected in experiments with porcine brain tubulin, and therefore must be small. Experiments suggest that the cap must be less than Ϸ200 dimers (2-4). Presumably, the transition from rapid shortening to growth, termed ''rescue,'' occurs when the tubulin-GTP cap is reestablished.Interactions at the surfaces of adjacent dimers occur through discrete lateral and longitudinal noncovalent bonds. Thermodynamic studies of MT assembly are unable to qua...

Microtubule self-assembly is largely governed by the chemical kinetics and thermodynamics of tubulin-tubulin interactions. An important aspect of microtubule assembly is that hydrolysis of the beta-tubulin-associated GTP promotes protofilament curling. Protofilament curling presumably drives the transition from tip structures associated with growth (sheetlike projections and blunt ends) to those associated with shortening (rams' horns and frayed ends), and transitions between these structures have been proposed to be important for growth-shortening transitions. However, previous models for microtubule dynamic instability have not considered such structures or mechanics explicitly. Here we present a three-dimensional model that explicitly incorporates mechanical stress and strain within the microtubule lattice. First, we found that the model recapitulates three-dimensional tip structures and rates of assembly and disassembly for microtubules grown under standard conditions, and we propose that taxol may stabilize microtubule growth by reducing flexural rigidity. Second, in contrast to recent suggestions, it was determined that sheetlike tips are more likely to undergo catastrophe than blunt tips. Third, partial uncapping of the tubulin-GTP cap provides a possible mechanism for microtubule pause events. Finally, simulations of the binding and structural effects of XMAP215 produced the experimentally observed growth and shortening rates, and tip structure.

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