scholarly journals P granules extend the nuclear pore complex environment in the C. elegans germ line

2011 ◽  
Vol 192 (6) ◽  
pp. 939-948 ◽  
Author(s):  
Dustin L. Updike ◽  
Stephanie J. Hachey ◽  
Jeremy Kreher ◽  
Susan Strome
Keyword(s):  
Nuclear Pore Complex ◽  
Germ Plasm ◽  
Germ Line ◽  
Hydrophobic Interactions ◽  
Nuclear Periphery ◽  
Size Exclusion ◽  
Nuclear Pore ◽  
Complex Environment ◽  
P Granules ◽  
C Elegans

The immortal and totipotent properties of the germ line depend on determinants within the germ plasm. A common characteristic of germ plasm across phyla is the presence of germ granules, including P granules in Caenorhabditis elegans, which are typically associated with the nuclear periphery. In C. elegans, nuclear pore complex (NPC)–like FG repeat domains are found in the VASA-related P-granule proteins GLH-1, GLH-2, and GLH-4 and other P-granule components. We demonstrate that P granules, like NPCs, are held together by weak hydrophobic interactions and establish a size-exclusion barrier. Our analysis of intestine-expressed proteins revealed that GLH-1 and its FG domain are not sufficient to form granules, but require factors like PGL-1 to nucleate the localized concentration of GLH proteins. GLH-1 is necessary but not sufficient for the perinuclear location of granules in the intestine. Our results suggest that P granules extend the NPC environment in the germ line and provide insights into the roles of the PGL and GLH family proteins.

scholarly journals Proximity labeling identifies LOTUS domain proteins that promote the formation of perinuclear germ granules in C. elegans

eLife ◽  
2021 ◽  
Vol 10 ◽  
Author(s):  
Ian F Price ◽  
Hannah L Hertz ◽  
Benjamin Pastore ◽  
Jillian Wagner ◽  
Wen Tang
Keyword(s):  
Germ Line ◽  
Nuclear Periphery ◽  
P Granules ◽  
C Elegans ◽  
Intrinsically Disordered ◽  
Intrinsically Disordered Regions ◽  
Germ Granules ◽  
Protein Components ◽  
Labeling Method ◽  
Disordered Regions

The germ line produces gametes that transmit genetic and epigenetic information to the next generation. Maintenance of germ cells and development of gametes require germ granules-well-conserved membraneless and RNA-rich organelles. The composition of germ granules is elusive owing to their dynamic nature and their exclusive expression in the germ line. Using C. elegans germ granule, called P granule, as a model system, we employed a proximity-based labeling method in combination with mass spectrometry to comprehensively define its protein components. This set of experiments identified over 200 proteins, many of which contain intrinsically disordered regions. An RNAi-based screen identified factors that are essential for P granule assembly, notably EGGD-1 and EGGD-2, two putative LOTUS-domain proteins. Loss of eggd-1 and eggd-2 results in separation of P granules from the nuclear envelope, germline atrophy and reduced fertility. We show that intrinsically disordered regions of EGGD-1 are required to anchor EGGD-1 to the nuclear periphery while its LOTUS domains are required to promote perinuclear localization of P granules. Together, our work expands the repertoire of P granule constituents and provides new insights into the role of LOTUS-domain proteins in germ granule organization.

scholarly journals Novel LOTUS-domain proteins are organizational hubs that recruit C. elegans Vasa to germ granules

eLife ◽  
2021 ◽  
Vol 10 ◽  
Author(s):  
Patricia Giselle Cipriani ◽  
Olivia Bay ◽  
John Zinno ◽  
Michelle Gutwein ◽  
Hin Hark Gan ◽  
...  
Keyword(s):  
Rna Processing ◽  
Germ Line ◽  
Temperature Sensitive ◽  
Developmental Transitions ◽  
P Granules ◽  
C Elegans ◽  
Intrinsically Disordered ◽  
Meiotic Progression ◽  
Intrinsically Disordered Regions ◽  
Granule Growth

We describe MIP-1 and MIP-2, novel paralogous C. elegans germ granule components that interact with the intrinsically disordered MEG-3 protein. These proteins promote P granule condensation, form granules independently of MEG-3 in the postembryonic germ line, and balance each other in regulating P granule growth and localization. MIP-1 and MIP-2 each contain two LOTUS domains and intrinsically disordered regions and form homo- and heterodimers. They bind and anchor the Vasa homolog GLH-1 within P granules and are jointly required for coalescence of MEG-3, GLH-1, and PGL proteins. Animals lacking MIP-1 and MIP-2 show temperature-sensitive embryonic lethality, sterility, and mortal germ lines. Germline phenotypes include defects in stem cell self-renewal, meiotic progression, and gamete differentiation. We propose that these proteins serve as scaffolds and organizing centers for ribonucleoprotein networks within P granules that help recruit and balance essential RNA processing machinery to regulate key developmental transitions in the germ line.

scholarly journals Telomere relocalization to the nuclear pore complex in response to replication stress

2020 ◽  
Author(s):  
Alexandra M Pinzaru ◽  
Noa Lamm ◽  
Mike al-Kareh ◽  
Eros Lazzerini-Denchi ◽  
Anthony J Cesare ◽  
...  
Keyword(s):  
Nuclear Pore Complex ◽  
Actin Polymerization ◽  
Nuclear Periphery ◽  
Tumor Development ◽  
Replication Stress ◽  
Nuclear Pore ◽  
Telomere Repeat ◽  
Telomere Binding Protein ◽  
The Impact ◽  
In Cells

AbstractMutations in the telomere binding protein, POT1 are associated with solid tumors and leukemias. POT1 alterations cause rapid telomere elongation, ATR kinase activation, telomere fragility, and accelerated tumor development. Here, we investigated the impact of mutant POT1 alleles through complementary genetic and proteomic approaches based on CRISPR-interference and biotin-based proximity labelling, respectively. These screens revealed that replication stress is a major vulnerability in cells expressing mutant POT1 and manifest in increased mitotic DNA synthesis (MiDAS) at telomeres. Our study also unveiled a role for the nuclear pore complex (NPC) in resolving replication defects at telomeres. Depletion of NPC subunits in the context of POT1 dysfunction increased DNA damage signaling and telomere fragility. Furthermore, we observed telomere repositioning to the nuclear periphery driven by nuclear F-actin polymerization in cells with POT1 mutations. In conclusion, our study establishes that relocalization of dysfunctional telomeres to the nuclear periphery is critical to preserve telomere repeat integrity.

Germline and developmental roles of the nuclear transport factor importin (α)3 in C. elegans

Development ◽  
2001 ◽  
Vol 128 (10) ◽  
pp. 1817-1830 ◽  
Author(s):  
K.G. Geles ◽  
S.A. Adam
Keyword(s):  
Nuclear Transport ◽  
Intracellular Localization ◽  
Nuclear Pore ◽  
P Granules ◽  
C Elegans ◽  
Cellular Events ◽  
Importin Α ◽  
Localization Signal ◽  
Transport Factors ◽  
Embryonic And Larval Development

The importin (α) family of transport factors mediates the nuclear import of classical nuclear localization signal-containing proteins. In order to understand how multiple importin (α) proteins are regulated both in individual cells and in a whole organism, the three importin (α) (ima) genes of Caenorhabditis elegans have been identified and studied. All three IMAs are expressed in the germline; however, only IMA-3 is expressed in the soma. RNA interference (RNAi) experiments demonstrate that IMA-3 is required for the progression of meiotic prophase I during oocyte development. Loss of IMA-3 expression leads also to a disruption of the nuclear pore complex accompanied by the mis-localization of P granules. A range of defects occurring in ima-3(RNAi) F(1) progeny further supports a role for IMA-3 during embryonic and larval development. The functional association of IMA-3 with distinct cellular events, its expression pattern and intracellular localization indicate that regulation of the nuclear transport machinery is involved in the control of developmental pathways.

scholarly journals Caenorhabditis elegans Nucleoporins Nup93 and Nup205 Determine the Limit of Nuclear Pore Complex Size Exclusion In Vivo

2003 ◽  
Vol 14 (12) ◽  
pp. 5104-5115 ◽  
Author(s):  
Vincent Galy ◽  
Iain W. Mattaj ◽  
Peter Askjaer
Keyword(s):  
Caenorhabditis Elegans ◽  
Nuclear Envelope ◽  
Protein Import ◽  
Chromatin Condensation ◽  
Size Exclusion ◽  
Nuclear Pore ◽  
Nuclear Pore Complexes ◽  
C Elegans ◽  
Pore Complexes

Nuclear pore complexes (NPCs) span the nuclear envelope and mediate communication between the nucleus and the cytoplasm. To obtain insight into the structure and function of NPCs of multicellular organisms, we have initiated an extensive analysis of Caenorhabditis elegans nucleoporins. Of 20 assigned C. elegans nucleoporin genes, 17 were found to be essential for embryonic development either alone or in combination. In several cases, depletion of nucleoporins by RNAi caused severe defects in nuclear appearance. More specifically, the C. elegans homologs of vertebrate Nup93 and Nup205 were each found to be required for normal NPC distribution in the nuclear envelope in vivo. Depletion of Nup93 or Nup205 caused a failure in nuclear exclusion of nonnuclear macromolecules of ∼70 kDa without preventing active nuclear protein import or the assembly of the nuclear envelope. The defects in NPC exclusion were accompanied by abnormal chromatin condensation and early embryonic arrest. Thus, the contribution to NPC structure of Nup93 and Nup205 is essential for establishment of normal NPC function and for cell viability.

scholarly journals Nuclear pore complex evolution: a trypanosome Mlp analogue functions in chromosomal segregation but lacks transcriptional barrier activity

2014 ◽  
Vol 25 (9) ◽  
pp. 1421-1436 ◽  
Author(s):  
Jennifer M. Holden ◽  
Ludek Koreny ◽  
Samson Obado ◽  
Alexander V. Ratushny ◽  
Wei-Ming Chen ◽  
...  
Keyword(s):  
Nuclear Pore Complex ◽  
Mitotic Spindle ◽  
Nuclear Periphery ◽  
Coiled Coil ◽  
Nuclear Lamina ◽  
Nucleocytoplasmic Transport ◽  
Nuclear Pore ◽  
Dependent Manner ◽  
Major Barrier ◽  
African Trypanosomes

The nuclear pore complex (NPC) has dual roles in nucleocytoplasmic transport and chromatin organization. In many eukaryotes the coiled-coil Mlp/Tpr proteins of the NPC nuclear basket have specific functions in interactions with chromatin and defining specialized regions of active transcription, whereas Mlp2 associates with the mitotic spindle/NPC in a cell cycle–dependent manner. We previously identified two putative Mlp-related proteins in African trypanosomes, TbNup110 and TbNup92, the latter of which associates with the spindle. We now provide evidence for independent ancestry for TbNup92/TbNup110 and Mlp/Tpr proteins. However, TbNup92 is required for correct chromosome segregation, with knockout cells exhibiting microaneuploidy and lowered fidelity of telomere segregation. Further, TbNup92 is intimately associated with the mitotic spindle and spindle anchor site but apparently has minimal roles in control of gene transcription, indicating that TbNup92 lacks major barrier activity. TbNup92 therefore acts as a functional analogue of Mlp/Tpr proteins, and, together with the lamina analogue NUP-1, represents a cohort of novel proteins operating at the nuclear periphery of trypanosomes, uncovering complex evolutionary trajectories for the NPC and nuclear lamina.

scholarly journals Germline maintenance through the multifaceted activities of GLH/Vasa in Caenorhabditis elegans P granules

2019 ◽  
Author(s):  
Elisabeth A. Marnik ◽  
J. Heath Fuqua ◽  
Catherine S. Sharp ◽  
Jesse D. Rochester ◽  
Emily L. Xu ◽  
...  
Keyword(s):  
Small Rna ◽  
Nuclear Periphery ◽  
Protein Translation ◽  
26S Proteasome ◽  
Mass Spectrometry Analysis ◽  
List Type ◽  
Helicase Activity ◽  
Critical Component ◽  
P Granules ◽  
C Elegans

SUMMARYVasa is a highly conserved member of the ATP-dependent DEAD box helicase family, a multipotency factor, and a critical component for the specification and maintenance of the germline. Its homologs have been shown to regulate translation, small RNA amplification, and serve as a molecular solvent for single-stranded RNA; however, the function of Vasa’s defining domains and what they interact with are unclear. To address this, 28 mutant alleles of the C. elegans Vasa homolog GLH-1 were generated in conserved motifs. Mutations in the flanking and helicase domains show that GLH-1 retains its association with P granules through its helicase activity and not through static interactions with other P-granule proteins. Changes outside of these domains retain GLH-1 in P granules but still compromise fertility, and removal of glycine-rich repeats progressively diminish P-granule wetting-like interactions at the nuclear periphery. A mutation that facilitates Vasa aggregation was previously leveraged in insects and mammals to identify the transient association of Vasa with piRNA amplifying Argonautes. This same mutation in GLH-1 also stimulates aggregation and association with Argonautes, suggesting that the transient amplifying complex is evolutionarily conserved even though the method of piRNA amplification in C. elegans is not. Mass spectrometry analysis of proteins that co-immunoprecipitate with wild type and mutant GLH-1 reveal an affinity for all three PCI (26S Proteasome Lid, COP9, eIF3) scaffolding complexes, which regulate protein turnover and translation, and a possible aversion for ribosomes and the 26S proteasome core. These results suggest that phase-separated P granules compartmentalize the cytoplasm to exclude large protein assemblies and emphasize the role of Vasa homologs in maintaining proteostasis. GRAPHICAL ABSTRACTHIGHLIGHTSGLH-1/Vasa helicase activity is required for germ granule association and the flanking domain is critical component of this helicase activity.GLH-1 and GLH-2 glycine-rich FG-repeats increase the coverage or wetting-like properties of germ granules at the nuclear periphery.Locked GLH-1 helicase domains increase association with Argonaute proteins, resembling small RNA transient amplifying complexes observed in insects and mammals.GLH-1 has an affinity for all three PCI (26S Proteasome Lid, COP9, eIF3) scaffolding complexes, emphasizing a role in protein translation and turnover.

scholarly journals Random sub-diffusion and capture of genes by the nuclear pore reduces dynamics and coordinates interchromosomal movement

2021 ◽  
Author(s):  
Michael Chas Sumner ◽  
Steven B. Torrisi ◽  
Donna Garvey Brickner ◽  
Jason H. Brickner
Keyword(s):  
Brownian Motion ◽  
Fractional Brownian Motion ◽  
Nuclear Pore Complex ◽  
Nuclear Periphery ◽  
Experimental Models ◽  
Nuclear Pore ◽  
Dynamic Tracking ◽  
Diffusive Movement ◽  
Chromosomal Loci

ABSTRACTHundreds of genes interact with the yeast nuclear pore complex (NPC), localizing at the nuclear periphery and clustering with co-regulated genes. Dynamic tracking of peripheral genes shows that they cycle on and off the NPC and that interaction with the NPC slows their sub-diffusive movement. Furthermore, NPC-dependent inter-chromosomal clustering leads to coordinated movement of pairs of loci separated by hundreds of nanometers. We developed Fractional Brownian Motion simulations for chromosomal loci in the nucleoplasm and interacting with NPCs. These simulations predict the rate and nature of random sub-diffusion during repositioning from nucleoplasm to periphery and match measurements from two different experimental models, arguing that recruitment to the nuclear periphery is due to random subdiffusion, collision, and capture by NPCs. Finally, the simulations do not lead to inter-chromosomal clustering or coordinated movement, suggesting that interaction with the NPC is necessary, but not sufficient, to cause clustering.

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