Evidence of Non-microtubule Spindle Forces in Mesostoma ehrenbergii Spermatocytes

Abstract

We tested conclusions reached in previous experiments in which Mesostoma spermatocyte chromosomes moved rapidly to a pole in the absence of microtubules: after 10 µM nocodazole (NOC) depolymerized metaphase spindle microtubules, kinetochores from each of the 3 bivalents detached from the same pole and rapidly moved to the other pole, at speeds averaging 37.7 µm/min. with some as high as 100 µm/min. We concluded that these very fast movements were due to nonmicrotubule forces arising from a spindle matrix. However, since the chromosomes stretch out before detaching, there is tension in the chromosomes from the stretch. Thus the movements of detached kinetochores conceivably might be due to recoil from the tension, though we argued against this possibility (Fegaras and Forer, 2018a). In this article we test whether recoil causes the movements. We cut bivalents into 2 pieces, using a femtosecond laser, before addition of NOC. When 1 bivalent was severed, all kinetochores moved to one pole in 12/15 cells; when 2 bivalents were severed, all kinetochores moved to one pole in 4/6 cells; and when all 3 bivalents were severed all kinetochores moved to one pole in 3/9 cells. The bivalent "halves" moved rapidly, with average speeds of 47 µm/min, velocities that are not significantly different from those in cells without any laser-cut bivalents (p > 0.05). Since kinetochores move at the same speeds whether they are part of bivalents or not, NOC-induced chromosome movements are not due to recoil from tension along the full-length bivalent, strongly supporting the idea that non-microtubule forces move chromosomes in Mesostoma spermatocytes.

Blocking Protein Phosphatase 1 [PP1] Prevents Loss of Tether Elasticity in Anaphase Crane-Fly Spermatocytes

“…After depolymerising spindle microtubules with nocodazole, each of the three bivalents stretches out between the poles and then after some minutes all three kinetochores at one pole detach from that pole and move rapidly to the other pole (Fegaras and Forer, 2018 ). Disabling any one of the three tethers, however, uncouples the coordination, such that after treatment with nocodazole different bivalents move to different poles, or do not move at all (Fegaras-Arch et al, 2020 ). Somehow tethers are necessary to coordinate the movements of these three different chromosomes.…”

Elastic tethers remain functional during anaphase arrest in partially-lysed crane-fly spermatocytes: a possible approach for studying mitotic tethers

“…The original experiments (LaFountain et al, 2002) were done using crane-fly spermatocytes but subsequent experiments using a laser microbeam to cut chromosome arms found the same phenomena in a wide variety of animal cells ranging from aquatic flatworms to insects to spiders to marsupials to humans (Forer et al, 2017). There is some evidence that tethers are involved in coordinating movements between chromosomes (Sheykhani et al, 2017; Forer and Berns, 2020; Fegaras-Arch et al, 2020).…”