Biology subjects

McNally, F.

Publications and source records attributed to McNally, F..

3 recordsLinked to original sources

bioRxiv2024

Mechanisms of Meiotic Spindle Initiation in Caenorhabditis elegans Oocytes

In many animal species, the oocyte meiotic spindle, which is required for chromosome segregation, forms without centrosomes. In some systems, Ran-GEF on chromatin initiates spindle assembly. We found that in C. elegans oocytes, endogenously-tagged Ran-GEF dissociates from chromatin during spindle assembly but re-associates during meiotic anaphase. Meiotic spindle assembly occurred after auxin-induced degradation of Ran-GEF but anaphase I was faster than controls and extrusion of the first polar body frequently failed. In search of a possible alternative pathway for spindle assembly, we found that soluble tubulin concentrates in the nuclear volume during germinal vesicle breakdown. We found that the concentration of soluble tubulin in the metaphase spindle region is enclosed by ER sheets which exclude cytoplasmic organelles including mitochondria and yolk granules. Measurement of the volume occupied by yolk granules and mitochondria indicated that volume exclusion would be sufficient to explain the concentration of tubulin in the spindle volume. We suggest that this concentration of soluble tubulin may be a redundant mechanism promoting spindle assembly near chromosomes.

cell biology↗

bioRxiv2019

Spherical Spindle Shape Promotes Perpendicular Cortical Orientation by Preventing Isometric Cortical Pulling on both Spindle Poles during C. elegans Female Meiosis

Meiotic spindles are positioned perpendicular to the oocyte cortex to facilitate segregation of chromosomes into a large egg and a tiny polar body. In C. elegans, spindles are initially ellipsoid and parallel to the cortex before shortening to a spherical shape and rotating to the perpendicular orientation by dynein-driven cortical pulling. The mechanistic connection between spindle shape and rotation has remained elusive. Here we used mutants of the microtubule-severing protein katanin to manipulate spindle shape without eliminating cortical pulling. In a katanin mutant, spindles remained ellipsoid, had pointed poles and became trapped in either a diagonal or a parallel orientation. Results indicated that astral microtubules emanating from both spindle poles initially engage in cortical pulling until microtubules emanating from one pole detach from the cortex allowing pivoting of the spindle. The lower viscous drag experienced by spherical spindles prevented recapture of the cortex by astral microtubules emanating from the detached pole. In addition, maximizing contact between pole dynein and cortical dynein stabilizes round poles in a perpendicular orientation. Spherical spindle shape can thus promote perpendicular orientation by two distinct mechanisms.

cell biology↗

bioRxiv2017

Structural Basis for Katanin Self-Assembly

The reorganization of microtubules in mitosis, meiosis and development requires the microtubule-severing activity of katanin. Katanin is composed of a AAA ATPase subunit and a regulatory subunit. Microtubule severing requires ATP hydrolysis by katanins conserved AAA ATPase domains. Whereas other AAA ATPases form stable hexamers, we show that wild-type katanin only forms heterodimers and heterotetramers. Heterododecamers were only observed for an ATP hydrolysis deficient mutant in the presence of ATP, suggesting an auto-inhibition mechanism that prevents oligomerization. X-ray structures of katanins AAA ATPase in monomeric nucleotide-free and pseudo-oligomeric ADP-bound states reveal conformational changes in AAA subdomains and N and C-terminal expansion segments that explain this auto-inhibition of assembly. These data lead to a model in which self-inhibited heterodimers bind to a microtubule, then transition into an assembly-competent conformation upon ATP binding. Microtubule-bound heterododecamers then promote tubulin extraction from the microtubule prior to oligomer dissociation.

biochemistry↗