Electrostatic Considerations in Mitosis

1.4 Summary In light of the large body of experimental information regardingmitosis, the complexity and lack of unity of models for the various events and motions gives, at least to this observer, reason to believe that approaching mitosis primarily within the molecular biology paradigm is flawed. This book offers a different approach based on nanoscale electrostatic interactions. It seems clear that cellular electrostatics involves more than the traditional thinking regarding counterion screening of electric fields and the resulting unimportance within cells of the second most powerful force in nature. The reality may be that the evidence suggests otherwise, and that the resulting enhanced electrostatic interactions are more robust and act over greater distances than previously thought. One aspect of this is the ability of microtubules to extend the reach of electrostatic force over cellular distances; another lies in the reduced counterion screening and dielectric constant of the cytosol between charged protein surfaces. High pHi during prophase favors spindle assembly. This includes greater electrostatic attractive forces between tubulin dimers as well as increased repulsive electrostatic interactions driving poleward movements of forming half-spindles. Changes in microtubule dynamics are integral to changes in the motions of chromosomes during mitosis. These changes in mi- crotubule dynamics can be attributed to an associated change in intracellular pH (pHi) during mitosis. In particular, a decrease in pHi during mitosis may act as a master clock controlling microtubule disassembly/assembly probability ratios by altering the electrostatic interactions of tubulin dimers. This, in turn, could determine the timing and dynamics of post-attachment mitotic chromosome motions. The possible electrostatic consequences of a subsequently decreasing pHi on mitotic motions and events will be discussed in chapters to follow. 3.5 Summary Given the known net negative charge at the plus ends of microtubules and the presence of highly basic molecules in kinetochores it is difficult to imagine there not being an attractive electrostatic poleward-directed force between the plus ends of kinetochore microtubules and kinetochores. Calculations of electrostatic force magnitudes for penetrating and non-penetrating microtubules within critical separations show that nanoscale electrostatic interactions are able to account for poleward-directed force production at both kinetochores and poles. The calculated maximum force per microtubule falls within the experimental range, and represents a successful ab initio derivation of the magnitude of this force. A simulation (see Section 4.4) supports this calculation. In agreement with experiment [Nicklas, 1988], the calculations given in this chapter satisfy the requirement that the maximum tension force per microtubule be the same for all mitotic chromosome attachments. The approximate equality in the calculations for electrostatic force generation by non-penetrating or penetrating kinetochore microtubules at both poles and kinetochores, as well as parity between overall force generation at poles and kinetochores demonstrated here, is to be expected of any dynamical mechanism for poleward force generation. Models for chromosome motility must account for these parities. Models for chromosome motility should also address the efficiency with which spindle microtubules maintain coupling to kinetochores and poles throughout the various movements during mitosis. Electrostatic force at nanometer distances between the free ends of a kinetochore microtubule and a kinetochore or centrosome matrix has this property. Consistent with observation, spindle microtubules depolymerize while generating force at kinetochores and poles. Given the electrostatic nature of tubulin microtubule subunits, this can be understood in terms of the large electric field (and therefore force) gradients in critical distances within vicinal cytosol outside as well as across the boundaries of kinetochores and cen- trosome matrices. 5.6 Summary Post-attachment chromosome motions during prometaphase and metaphase can be explained by statistical fluctuations in nanoscale electrostatic microtubule antipoleward assembly forces acting between microtubules and chromosome arms, combined with similar fluctuations in nanoscale electrostatic microtubule poleward disassembly forces acting at kinetochores and spindle poles. The different motions throughout prometaphase and metaphase may be understood as an increase in the microtubule disassembly to assembly probability ratio. It seems reasonable to assume that the shift from the dominance of microtubule growth during prophase, and to a lesser extent during prometaphase, to a parity between microtubule polymerization and depolymerization during metaphase chromosome oscillations could be attributed to the gradual downward pHi shift during mitosis that is observed in many cell types. Evidence for a continuing decrease in pHi and an increasing microtubule disassembly to assembly probability ratio is also seen in increased kinetochore tension just prior to anaphase. This increased tension has a possible simple interpretation in terms of the greater magnitude of poleward electrostatic disassembly forces at sister kinetochores and poles relative to antipoleward assembly forces, as well as an increased mutual repulsion of sister kinetochores due to a greater expression of kinetochore positive charge, both due to decreased pHi. There does not appear to be consensus on a model for the generation of poleward force at cell poles. Experimental observations regarding the microtubule disassembly force at poles, with an associated microtubule flux, can be consistently explained in terms of the same nanoscale electrostatic force mechanism as that operating at kinetochores.... The observed intracellular increase in [Ca2+] that occurs at the onset of anaphase-A further increases the probability for microtubule disassembly, effectively dwarfing antipoleward nanoscale microtubule assembly forces from both poles. The sudden dominance of poleward nanoscale microtubule disassembly forces added to a greater mutual repulsion of positively charged sister kinetochores, with both due to a decreased pHi could be integral in the initial separation of sister chromatids. Once this separation is effected, anaphase-A motion would result from the predominance of electrostatic microtubule disassembly forces at kinetochores and poles.