The homotypic fusion and protein sorting (HOPS) complex is a multi-subunit complex conserved from yeast to mammals that regulates late endosome and lysosome fusion. However, little is known about how the HOPS complex is recruited to lysosomes in mammalian cells. Here, we report that the small GTPase Arl8b, but not Rab7 (also known as RAB7A), is essential for membrane localization of the human (h)Vps41 subunit of the HOPS complex. Assembly of the core HOPS subunits to Arl8b- and hVps41-positive lysosomes is guided by their subunit–subunit interactions. RNA interference (RNAi)-mediated depletion of hVps41 resulted in the impaired degradation of EGFR that was rescued upon expression of wild-type but not an Arl8b-binding-defective mutant of hVps41, suggesting that Arl8b-dependent lysosomal localization of hVps41 is required for its endocytic function. Furthermore, we have also identified that the Arl8b effector SKIP (also known as PLEKHM2) interacts with and recruits HOPS subunits to Arl8b and kinesin-positive peripheral lysosomes. Accordingly, RNAi-mediated depletion of SKIP impaired lysosomal trafficking and degradation of EGFR. These findings reveal that Arl8b regulates the association of the human HOPS complex with lysosomal membranes, which is crucial for the function of this tethering complex in endocytic degradation.

Highlights d Feedback regulation between Hop1, Zip1, and Pch2 influences the synaptonemal complex d Pch2 and Hop1 collaborate to mediate general meiotic prophase checkpoint function d Pch2 has activating and silencing roles in meiotic prophase checkpoint function d Synaptonemal complex polymerization dictates switch-like behavior of Pch2 function

One of the defining features of sexual reproduction is the recombination events that take place during meiosis I. Recombination is both evolutionarily advantageous, but also mechanistically necessary to form the crossovers that link homologous chromosomes. Meiotic recombination is initiated through the placement of programmed double-strand DNA breaks (DSBs) mediated by the protein Spo11. The timing, number, and physical placement of DSBs are carefully controlled through a variety of protein machinery. Previous work has implicated Mer2(IHO1 in mammals) to be involved in both the placement of breaks, and their timing. In this study we use a combination of protein biochemistry and biophysics to extensively characterise various roles of the Mer2. We gain further insights into the details of Mer2 interaction with the PHD protein Spp1, reveal that Mer2 is a novel nucleosome binder, and suggest how Mer2’s interaction with the HORMA domain protein Hop1 (HORMAD1/2 in mammals) is controlled.

In meiosis, DNA double strand break (DSB) formation by Spo11 initiates recombination and enables chromosome segregation. Numerous factors are required for Spo11 activity, and couple the DSB machinery to the development of a meiosis-specific “axis-tethered loop” chromosome organization. Through in vitro reconstitution and budding yeast genetics we here provide architectural insight into the DSB machinery by focussing on a foundational DSB factor, Mer2. We characterise the interaction of Mer2 with the histone reader Spp1, and show that Mer2 directly associates to nucleosomes, likely highlighting a contribution of Mer2 to tethering DSB factors to chromatin. We reveal the biochemical basis of Mer2 association with Hop1, a HORMA domain-containing chromosomal axis factor. Finally, we identify a conserved region within Mer2 crucial for DSB activity, and show that this region of Mer2 interacts with the DSB factor Mre11. In combination with previous work, we establish Mer2 as a keystone of the DSB machinery by bridging key protein complexes involved in the initiation of meiotic recombination.