The KASH5 protein involved in meiotic chromosomal movements is a novel dynein activating adaptor
Curation statements for this article:-
Curated by eLife
Evaluation Summary:
This study contributes to our understanding of how a diverse and increasing number of activating adaptors allow the dynein motor protein to move a wide range of intracellular cargoes. Here the authors identify a transmembrane protein called KASH5 as the activating adaptor required for dynein to move meiotic chromosomes, a process that facilitates homolog pairing. Overall, the work is well done and will be of interest to the cell biology, cytoskeletal, and meiosis research communities.
(This preprint has been reviewed by eLife. We include the public reviews from the reviewers here; the authors also receive private feedback with suggested changes to the manuscript. Reviewer #1 and Reviewer #2 agreed to share their names with the authors.)
This article has been Reviewed by the following groups
Listed in
- Evaluated articles (eLife)
- Cell Biology (eLife)
Abstract
Dynein harnesses ATP hydrolysis to move cargo on microtubules in multiple biological contexts. Dynein meets a unique challenge in meiosis by moving chromosomes tethered to the nuclear envelope to facilitate homolog pairing essential for gametogenesis. Though processive dynein motility requires binding to an activating adaptor, the identity of the activating adaptor required for dynein to move meiotic chromosomes is unknown. We show that the meiosis-specific nuclear-envelope protein KASH5 is a dynein activating adaptor: KASH5 directly binds dynein using a mechanism conserved among activating adaptors and converts dynein into a processive motor. We map the dynein-binding surface of KASH5, identifying mutations that abrogate dynein binding in vitro and disrupt recruitment of the dynein machinery to the nuclear envelope in cultured cells and mouse spermatocytes in vivo. Our study identifies KASH5 as the first transmembrane dynein activating adaptor and provides molecular insights into how it activates dynein during meiosis.
Article activity feed
-
-
Evaluation Summary:
This study contributes to our understanding of how a diverse and increasing number of activating adaptors allow the dynein motor protein to move a wide range of intracellular cargoes. Here the authors identify a transmembrane protein called KASH5 as the activating adaptor required for dynein to move meiotic chromosomes, a process that facilitates homolog pairing. Overall, the work is well done and will be of interest to the cell biology, cytoskeletal, and meiosis research communities.
(This preprint has been reviewed by eLife. We include the public reviews from the reviewers here; the authors also receive private feedback with suggested changes to the manuscript. Reviewer #1 and Reviewer #2 agreed to share their names with the authors.)
-
Reviewer #1 (Public Review):
This manuscript identifies a meiosis-specific protein that recruits and activates the motility of the dynein-1 transport machinery at the nuclear envelope. In prophase I of meiosis, dynein moves chromosomes tethered to the nuclear envelope to expedite the search and pairing between homologous chromosomes. Previous studies have shown that dynein tethers to chromosomes via the LINC complex, which consists of a SUN protein and transmembrane KASH protein. KASH5 comprises all the known features of bona fide cargo adaptors of dynein. A previous study reported an association between dynein and KASH5, but it remained to be demonstrated whether KASH5 directly binds dynein and activates its processive motility. Overall, the work is well done and will be broadly interesting to the cell biology and biophysics readership.
-
Reviewer #2 (Public Review):
The manuscript by Agrawal et al. describes a molecular analysis of the meiosis-specific nesprin-family protein KASH5. KASH5 is known to interact with dynein and dynactin, but the biological function of this interaction remained obscure. The authors utilize biochemistry and single-molecule analysis to demonstrate that KASH5 acts as an activating adapter molecule that links the dynein motor to dynactin and activates processive dynein motility along microtubules. They map the interaction between KASH5 and the dynein LIC subunit to show that KASH5 binds to the dynein LIC in a manner that is very similar to previously reported dynein adapter proteins. They test multiple point mutations that are predicted to disrupt this interaction and find a subset that both disrupt the KASH5-dynein interaction in vitro and in …
Reviewer #2 (Public Review):
The manuscript by Agrawal et al. describes a molecular analysis of the meiosis-specific nesprin-family protein KASH5. KASH5 is known to interact with dynein and dynactin, but the biological function of this interaction remained obscure. The authors utilize biochemistry and single-molecule analysis to demonstrate that KASH5 acts as an activating adapter molecule that links the dynein motor to dynactin and activates processive dynein motility along microtubules. They map the interaction between KASH5 and the dynein LIC subunit to show that KASH5 binds to the dynein LIC in a manner that is very similar to previously reported dynein adapter proteins. They test multiple point mutations that are predicted to disrupt this interaction and find a subset that both disrupt the KASH5-dynein interaction in vitro and in vivo. They finally test these mutations in meiotic mouse spermatocytes and show that some of the mutations predicted to disrupt the KASH5-dynein interaction also disrupt dynein recruitment to telomeres. This disruption is, however, more mild than the phenotypes caused by these mutations in non-meiotic cells. Overall, I found the manuscript to be highly accessible and the conclusions are well-supported by the data. The discovery of KASH5 as a dynein activating adapter is very interesting for the field and represents a novel class of transmembrane activating adapters. I do not have substantial reservations about the data, which I find is high quality and believable.
One of the more interesting aspects of the work that I think warrants more discussion from the authors is the stoichiometry of the KASH5-dynein complex. The SUN-Nesprin complexes are typically represented as trimers, a model based on the crystal structure of SUN2 (Sosa et al. Cell, 2012). In this work, the authors present evidence that KASH5 alone is a dimer and that the stoichiometry of the KASH5-LIC1 complex is 2:1. It is currently unclear to me how a KASH5 dimer might interact with a dynein dimer containing two LIC subunits. The authors mention a prior study that found a similar stoichiometry in the crystal structure for the well-studied adapter proteins BicD2 and LIC1 (Lee et al. Nat. Comm. 2020). However, that study presented evidence that the 2:1 stoichiometry observed in the crystal was likely a crystal packing artifact and that BicD2 bound to two LIC1 peptides in solution. This is something the authors of the current manuscript should mention and clarify in their discussion of these results. Further, it is interestingly unclear to me how a trimeric complex of SUN1-KASH5 might interact with the dimeric dynein complex. It is also unclear how a dimeric KASH5 would interact with a trimeric SUN1. The authors might consider expanding on their thoughts on the topic of the structural organization of the SUN-Nesprin-Dynein complexes in vivo, which would no doubt provide stimulating ideas for further experimentation.
A weakness of the paper is the somewhat mild phenotype observed in mouse spermatocytes expressing KASH5 mutants. While these same mutants had stronger phenotypes in non-meiotic cells, it is unclear why the phenotypes are more mild in the biologically relevant cell type. The authors' explanation of endogenous wild-type KASH5 blunting these effects seems plausible, but it would be nice to confirm. The authors do not try combining point mutations into a single construct, which may have stronger effects on dynein binding or titration of expression to higher levels to outcompete the endogenous KASH5 protein. Alternatively, it may be possible to knock down expression of the endogenous KASH5 protein using siRNA.
-
Reviewer #3 (Public Review):
This study contributes to understanding how a diverse and increasing number of activating adaptors allow dynein to move a wide range of intracellular cargoes. Here the authors identify a transmembrane protein called KASH5 as the activating adaptor required for dynein to move meiotic chromosomes, a process that facilitates homolog pairing.
It is an interesting study that combines in vitro reconstitutions with cellular measurements to validate KASH5 as the new dynein activating adaptor with some properties distinct from known adaptors. My main criticism, which should not require additional data collection but additional data analysis, is to quantify run-lengths and some measure of the number of processive events in the in vitro reconstitutions, as these are parameters that are as important as the velocity …
Reviewer #3 (Public Review):
This study contributes to understanding how a diverse and increasing number of activating adaptors allow dynein to move a wide range of intracellular cargoes. Here the authors identify a transmembrane protein called KASH5 as the activating adaptor required for dynein to move meiotic chromosomes, a process that facilitates homolog pairing.
It is an interesting study that combines in vitro reconstitutions with cellular measurements to validate KASH5 as the new dynein activating adaptor with some properties distinct from known adaptors. My main criticism, which should not require additional data collection but additional data analysis, is to quantify run-lengths and some measure of the number of processive events in the in vitro reconstitutions, as these are parameters that are as important as the velocity measurements that are shown.
-
