Integrity of Narrow Epithelial Tubes in the C. elegans Excretory System Requires a Transient Luminal Matrix

Gill, Hasreet K.; Cohen, Jozef; Ayala-Figueroa, Jesus; Forman-Rubinsky, Rachel; Poggioli, Corey; Bickard, Kevin; Parry, Jean M.; Pu, Pu; Hall, David H.; Sundaram, Meera V.

doi:10.1371/journal.pgen.1006205

Cited by 56 publications

(139 citation statements)

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“…Ultrastructural analysis of exc-2 mutants found visible fibrous material in the lumen (Fig. 2B’), which has also been seen only in other excretory system mutants affecting apical cytoskeleton proteins: sma-1 (encoding β H spectrin, erm-1 (ezrin-moesin-radixin homologue), and the excretory duct cell gene let-653 (mucin) (MCKEOWN et al 1998; BUECHNER et al 1999; KHAN et al 2013; GILL et al 2016). Future studies may show whether the fibrous material represents proteins or other material normally anchored to the membrane directly or indirectly by these cytoskeletal proteins.…”

Section: Discussionmentioning

confidence: 77%

“…In genetic screens, mutations in nine “ exc ” genes were discovered to affect canal structure to allow fluid-filled cysts to accumulate during canal extension during late embryogenesis and early first larval stage (BUECHNER et al 1999). Other studies found similar mutations that affect additional tubular structures in the nematode, including the seamed single-cell excretory duct cell (JONES and BAILLIE 1995; MANCUSO et al 2012; GILL et al 2016; PU et al 2017) and the multicellular intestine (BOSSINGER et al 2004; ZHANG et al 2011; CARBERRY et al 2012; KHAN et al 2013; ZHU et al 2015; GEISLER et al 2016). In the canal cell, proteins implementing tubule structure comprise apical cytoskeletal elements (MCKEOWN et al 1998; PRAITIS et al 2005; KHAN et al 2013; KOLOTUEV et al 2013; SHAYE and GREENWALD 2015), vesicular trafficking and exocyst proteins (TONG and BUECHNER 2008; MATTINGLY and BUECHNER 2011; ARMENTI et al 2014; LANT et al 2015; GRUSSENDORF et al 2016), and ion and lipid transporters (BERRY et al 2003; KHAN et al 2013), among others.…”

Section: Introductionmentioning

confidence: 70%

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Intermediate filaments EXC-2 and IFA-4 Maintain Luminal Structure of the Tubular Excretory Canals inCaenorhabditis elegans

Al-Hashimi

Hall

Ackley

et al. 2018

Preprint

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C. elegans excretory canals form a useful model for understanding formation of narrow 33 tubes. exc-2 mutants start to form normal canals that then swell into fluid-filled cysts. We show 34 that exc-2 encodes a large intermediate filament (IF) protein previously not thought to be located 35 in the canals. EXC-2 is located at the apical (luminal) membrane, binds to another IF protein, 36 and appears to be one of three IF proteins that form a flexible meshwork to maintain the thin 37 canal diameter. This work provides a genetically useful model for understanding the interactions 38 of IF proteins with other cytoskeletal elements to regulate tube size and growth. 39 3 ABSTRACT 40 The excretory canals of Caenorhabditis elegans are a model for understanding the maintenance 41 of apical morphology in narrow single-celled tubes. Light and electron microscopy shows that 42 mutants in exc-2 start to form canals normally, but these swell to develop large fluid-filled cysts 43 that lack a complete terminal web at the apical surface, and accumulate filamentous material in 44 the canal lumen. Here, whole-genome sequencing and gene rescue show that exc-2 encodes 45 intermediate filament protein IFC-2. EXC-2/IFC-2 protein, fluorescently tagged via 46 CRISPR/Cas9, is located at the apical surface of the canals independently of other intermediate 47 filament proteins. EXC-2 is also located in several other tissues, though the tagged isoforms are 48 not seen in the larger intestinal tube. Tagged EXC-2 binds via pulldown to intermediate filament 49 protein IFA-4, which is also shown to line the canal apical surface. Overexpression of either 50 protein results in narrow but shortened canals. These results are consistent with a model 51 whereby three intermediate filaments in the canals, EXC-2, IFA-4, and IFB-1, restrain swelling 52 of narrow tubules in concert with actin filaments that guide the extension and direction of tubule 53 outgrowth, while allowing the tube to bend as the animal moves. 54 93 While most of the original exc genes have been cloned (GRUSSENDORF et al. 2016; 94 SUNDARAM and BUECHNER 2016), mutations in the exc-2 gene cause particularly severe canal 95 defects. In these mutants, the canal length is shortened by over half, the animals accumulate 96 multiple cysts in the canals, and are sensitive to growth at low osmolarity (BUECHNER et al. 97 1999). Four alleles of this gene were discovered in the original screen, which suggested that it 98 encodes a large protein. Here we report that exc-2 encodes the intermediate filament IFC-2, and 99 additionally found that mutations in the ifa-4 intermediate filament gene also cause cystic canal 100 6 defects similar to those of exc-2 mutants. Overexpression of either exc-2 or ifa-4 results in 101 shortened canals with small or no cysts. EXC-2 and IFA-4 proteins bind to each other and are 102 located at the apical membrane of the canals. The position of EXC-2 at the apical membrane 103 occurs independently of IFB-1 and IFA-4 function in the canals. These result...

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Section: Discussionmentioning

confidence: 77%

Section: Introductionmentioning

confidence: 70%

Intermediate filaments EXC-2 and IFA-4 Maintain Luminal Structure of the Tubular Excretory Canals inCaenorhabditis elegans

Al-Hashimi

Hall

Ackley

et al. 2018

Preprint

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“…Notably, the lpr-1 mutant phenotype strongly resembles that caused by loss of the early aECM component LET-653 (Gill et al 2016). However, lpr-1 loss did not disrupt LET-653 secretion or localization in the duct (Gill et al 2016). Furthermore, LET-653 OE could not suppress lpr-1 mutant defects ( Figure S3), arguing against the model that let-653 is the key target upregulated by Ras signaling.…”

Section: Relationship Between Lpr-1 and Egf-ras-erk Signalingmentioning

confidence: 91%

“…Instead, we favor models where both LPR-1 and EGF signaling affect membrane lipids and/or the apical extracellular matrix (aECM), which are needed for lumen growth and maintenance. Notably, the lpr-1 mutant phenotype strongly resembles that caused by loss of the early aECM component LET-653 (Gill et al 2016). However, lpr-1 loss did not disrupt LET-653 secretion or localization in the duct (Gill et al 2016).…”

Section: Relationship Between Lpr-1 and Egf-ras-erk Signalingmentioning

confidence: 95%

The Lipocalin LPR-1 Cooperates with LIN-3/EGF Signaling To Maintain Narrow Tube Integrity in Caenorhabditis elegans

Pu¹,

Stone²,

Burdick³

et al. 2017

Genetics

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Lipocalins are secreted cup-shaped glycoproteins that bind sterols, fatty acids, and other lipophilic molecules. Lipocalins have been implicated in a wide array of processes related to lipophilic cargo transport, sequestration, and signaling, and several are used as biomarkers for human disease, but the functions of most lipocalins remain poorly understood. Here we show that the Caenorhabditis elegans lipocalin LPR-1 is required to maintain apical membrane integrity and a continuous lumen in two narrow unicellular tubes, the excretory duct and pore, during a period of rapid lumen elongation. LPR-1 fusion protein is expressed by the duct and pore and accumulates both intracellularly and in apical extracellular compartments, but it can also function cell nonautonomously when provided from outside of the excretory system. lpr-1 mutant defects can be rescued by increased signaling through the epidermal growth factor (EGF)-Ras-extracellular signal regulated kinase (ERK) pathway, which promotes the more elongated duct vs. less elongated pore tube fate. Spatial and temporal rescue experiments indicate that Ras signaling acts within the duct and pore tubes during or prior to cell fate determination to bypass the requirement for LPR-1. lpr-1 mutations did not disrupt LIN-3/EGF-dependent duct-fate specification, prevent functioning of any specific LIN-3/EGF isoform, or alter LET-23/EGFR localization, and reduced signaling did not phenocopy or enhance lpr-1 mutant defects. These data suggest that LPR-1 protects lumen integrity through a LIN-3/EGFindependent mechanism, but that increased signaling upregulates some target(s) that can compensate for lpr-1 absence.

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“…Red, duct cytoplasmic marker lin-48pro::mRFP . Green, luminal marker let-653pro::LET-653::GFP [ 100 ]. Scale bars.…”

Section: Figurementioning

confidence: 99%

Auto-fusion and the shaping of neurons and tubes

Soulavie

Sundaram

2016

Seminars in Cell & Developmental Biology

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Cells adopt specific shapes that are necessary for specific functions. For example, some neurons extend elaborate arborized dendrites that can contact multiple targets. Epithelial and endothelial cells can form tiny seamless unicellular tubes with an intracellular lumen. Recent advances showed that cells can auto-fuse to acquire those specific shapes. During auto-fusion, a cell merges two parts of its own plasma membrane. In contrast to cell-cell fusion or macropinocytic fission, which result in the merging or formation of two separate membrane bound compartments, auto-fusion preserves one compartment, but changes its shape. The discovery of auto-fusion in C. elegans was enabled by identification of specific protein fusogens, EFF-1 and AFF-1, that mediate cell-cell fusion. Phenotypic characterization of eff-1 and aff-1 mutants revealed that fusogen-mediated fusion of two parts of the same cell can be used to sculpt dendritic arbors, reconnect two parts of an axon after injury, or form a hollow unicellular tube. Similar auto-fusion events recently were detected in vertebrate cells, suggesting that auto-fusion could be a widely used mechanism for shaping neurons and tubes.

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Integrity of Narrow Epithelial Tubes in the C. elegans Excretory System Requires a Transient Luminal Matrix

Cited by 56 publications

References 102 publications

Intermediate filaments EXC-2 and IFA-4 Maintain Luminal Structure of the Tubular Excretory Canals inCaenorhabditis elegans

Intermediate filaments EXC-2 and IFA-4 Maintain Luminal Structure of the Tubular Excretory Canals inCaenorhabditis elegans

The Lipocalin LPR-1 Cooperates with LIN-3/EGF Signaling To Maintain Narrow Tube Integrity in Caenorhabditis elegans

Auto-fusion and the shaping of neurons and tubes

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