The meiotic LINC complex component KASH5 is an activating adaptor for cytoplasmic dynein

Abstract

Cytoplasmic dynein-driven movement of chromosomes during prophase I of meiosis is essential for synapsis and genetic exchange. Dynein connects to chromosome telomeres via KASH5 and SUN1/2, which span the outer and inner nuclear envelopes, respectively. Here, we show that KASH5 promotes dynein motility in vitro, and cytosolic KASH5 inhibits dynein’s interphase functions. KASH5 interacts with either dynein light intermediate chain (DYNC1LI1 or DYNC1LI2) via a conserved linker-helix in the LIC C-terminal, and this region is also needed for dynein’s recruitment to other cellular membranes. KASH5’s N-terminal EF-hands are essential, as the interaction with dynein is disrupted by mutation of key calcium-binding residues, although it is not regulated by cellular calcium levels. Dynein can be recruited to KASH5 at the nuclear envelope independently of dynactin, while LIS1 is essential for dynactin incorporation into the KASH5-dynein complex. Altogether, we show that the trans-membrane protein KASH5 is an activating adaptor for dynein, and shed light on the hierarchy of assembly of KASH5-dynein-dynactin complexes.

Molecular mechanism of dynein-dynactin activation by JIP3 and LIS1

“…In contrast addition of JIP3 1-185 resulted in multiple long distance runs of TMR-dynein (Figure 1C, D). The mean frequency over three independent experiments was 1.15 ±0.45 (SD) events/μm microtubule/min, which is very similar to that of the dynein adaptor HOOK3 collected under equivalent experimental conditions (Garner et al 2022). The mean velocity of the complexes was 1.00 ±0.2 μm/s (Figure 1E) which is also comparable to the velocity of HOOK3 complexes (Garner et al 2022).…”

“…The mean frequency over three independent experiments was 1.15 ±0.45 (SD) events/μm microtubule/min, which is very similar to that of the dynein adaptor HOOK3 collected under equivalent experimental conditions (Garner et al 2022). The mean velocity of the complexes was 1.00 ±0.2 μm/s (Figure 1E) which is also comparable to the velocity of HOOK3 complexes (Garner et al 2022). Under the conditions used in our assays we determined a run length for JIP3 complexes of 3.8 μm (Figure 1F).…”

“…Significance was determined using ANOVA with Tukey’s multiple comparison (ns = not significant, * = p≤0.05). The data for HOOK3 (Garner et al 2022) was used for comparison. (E) Distribution of velocities from 767 processive events for dynein-dynactin-JIP3 1-185 along with mean velocity ±S.D.…”

Dynamic regulation of LINC complex composition and function across tissues and contexts

“…However, to date there is little evidence that disrupting Nesprin‐3 abrogates tissue‐level function, as it is dispensable in a zebrafish model [26] and for male fertility in mice [25]. By contrast, KASH5, like SUN1, is essential for meiosis by functioning as an adapter to dynein [27–30]. Last, Nesprin‐4, as discussed above, impacts the function of outer hair cells in the ear [13] and plays roles more generally in secretory epithelia, where it is highly expressed [31].…”

Meiotic Chromosome Structure, the Synaptonemal Complex, and Infertility

“…Interestingly, meiotic cells express meiosis-specific cohesin subunits (75,92,93,129,133), at least some of which have evolved to carry out meiosis-specific roles (14). The telomeres at both ends of each chromosome become tethered to the nuclear envelope by the meiotic telomere complex (37,152), where they are subjected to microtubule forces that drive their rapid movement throughout the nucleus (51,68,91). These rapid prophase movements facilitate interhomolog recombination searches from the approximately 200-400 programmed DNA double-strand breaks (DSBs) that are generated in each cell (8,95,127).…”