Sm protein down-regulation leads to defects in nuclear pore complex disassembly and distribution in C. elegans embryos

Abstract

Summary Nuclear pore complexes (NPCs) are large macromolecular structures embedded in the nuclear envelope (NE), where they facilitate exchange of molecules between the cytoplasm and the nucleoplasm. In most cell types, NPCs are evenly distributed around the NE. However, the mechanisms dictating NPC distribution are largely unknown. Here, we used the model organism C. elegans to identify genes that affect NPC distribution during early embryonic divisions. We found that down-regulation of the Sm proteins, which are core components of the spliceosome, but not down-regulation of other splicing factors, led to clustering of NPCs. Down-regulation of Sm proteins also led to incomplete disassembly of NPCs during mitosis, but had no effect on lamina disassembly, suggesting that the defect in NPC disassembly was not due to a general defect in nuclear envelope breakdown. We further found that these mitotic NPC remnants persisted on an ER membrane that juxtaposes the mitotic spindle. At the end of mitosis, the remnant NPCs moved toward the chromatin and the reforming NE, where they ultimately clustered by forming membrane stacks perforated by NPCs. Our results suggest a novel, splicing-independent, role for Sm proteins in NPC disassembly, and point to a possible link between NPC disassembly in mitosis and NPC distribution in the subsequent interphase.

Down-regulation of tricarboxylic acid (TCA) cycle genes blocks progression through the first mitotic division in Caenorhabditis elegans embryos

“…Recently we conducted an RNAi screen in C. elegans for genes that affect nuclear morphology by targeting genes that were reported to cause embryonic lethality when mutated or down-regulated by RNAi (9). In the course of these studies we uncovered an unusual phenotype caused by the down-regulation of ORF T20G5.2 that codes for CTS-1, an ortholog of the eukaryotic citrate synthase (Fig.…”

“…Animals from RNAi plates were dissected on poly-Llysine-coated slides and embryos were opened by the freeze-cracking method as described previously (9). Samples were fixed in −20°C cold methanol for 2 min and blocked in 5% (vol/vol) BSA in PBS for 30 min at 4°C before overnight incubation at 4°C with primary antibody diluted in 1% BSA.…”

“…We have previously conducted a visual screen in C. elegans embryos for genes that when down-regulated by RNAi lead to an abnormal nuclear morphology (9). Most genes whose inactivation affected early embryonic development did so without arresting cell cycle progression.…”

A role for post-transcriptional control of ER dynamics and function in C. elegans germline stem cell maintenance

“…Although the NE is contiguous with the rough ER, not all NE proteins are present on the cytoplasmic ER network. Therefore, we generated double-transgenic lines expressing FARL-11::GFP and mCherry:: SP12, which is a marker for ER (Joseph-Strauss et al, 2012), and examined the distribution patterns of both proteins in germlines and embryos. As shown in Fig.…”

Alternative Splicing Regulation of Cancer-Related Pathways inCaenorhabditis elegans: An In Vivo Model System with a Powerful Reverse Genetics Toolbox

“…Their synmuv B mutant phenotypes are dependent on Ras activity and lin-3 ligand binding. While the activities, interactions, and mutant phenotypes of most of these splicing-related Rb enhancers are not well understood, RNAi of some of the snr genes ( snr-1, snr-2, snr-4, snr-5, snr-6 , and snr-7 ) led to embryonic lethality and nuclear pore organization disruption [56]. The question of whether these phenotypes arise from disrupted functions that have no role in splicing regulation is still unanswered, but evidence is mounting supporting the idea that constitutive splicing and alternative splicing are major contributors in C. elegans to pathways that, when altered, lead to cancer in human tissues.…”