The Rho-GEF PIX-1 directs assembly or stability of lateral attachment structures between muscle cells

Moody, Jasmine C.; Qadota, Hiroshi; Reedy, April R.; Okafor, C. Denise; Shanmugan, Niveda; Matsunaga, Yohei; Christian, Courtney J.; Ortlund, Eric A.; Benian, Guy M.

doi:10.1038/s41467-020-18852-4

Cited by 7 publications

(39 citation statements)

References 53 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…In addition, the effect of PAT-4 I432F overexpression was limited to muscle cell boundaries and dense bodies (somewhat), but not to M-lines. Our previous study suggests that the adhesion plaques at muscle cell boundaries are more dynamic than dense bodies or M-lines (Moody et al 2020): PIX-1 is a Rac GEF that is localized to all three integrin adhesion structures (M-lines, dense bodies and boundaries), but is only required at muscle cell boundaries. Either loss of function or overexpression of PIX-1 results in the same cell boundary defect (Moody et al 2020), suggesting that cell boundary structures require rapid turn over of component proteins, and is more dynamic than M-lines and dense bodies.…”

Section: Discussionmentioning

confidence: 99%

“…Our previous study suggests that the adhesion plaques at muscle cell boundaries are more dynamic than dense bodies or M-lines (Moody et al 2020): PIX-1 is a Rac GEF that is localized to all three integrin adhesion structures (M-lines, dense bodies and boundaries), but is only required at muscle cell boundaries. Either loss of function or overexpression of PIX-1 results in the same cell boundary defect (Moody et al 2020), suggesting that cell boundary structures require rapid turn over of component proteins, and is more dynamic than M-lines and dense bodies. In the future, we might be able to test this hypothesis by measuring and comparing protein exchange rates at the cell boundary, dense bodies and M-lines by, for example, FRAP analysis.…”

Section: Discussionmentioning

confidence: 99%

“…The membrane-proximal regions of the dense bodies and M-lines consist of integrin and associated proteins (Moerman and Williams 2006; Gieseler et al 2017). Integrin-associated proteins are also used at the adhesion plaques that lie between adjacent muscle cells, and are likely involved in transmitting forces laterally (Moody et al 2020). The cytoplasmic tail of β-integrin (PAT-3) is associated with four conserved proteins: UNC-112 (kindlin), PAT-4 (integrin-linked kinase, ILK), PAT-6 (α-parvin), and UNC-97 (PINCH).…”

Section: Introductionmentioning

confidence: 99%

See 2 more Smart Citations

Genetic Analysis Suggests a Surface of PAT-4 (ILK) that Interacts with UNC-112 (kindlin)

Benian

Qadota

McPherson

et al. 2022

Preprint

Self Cite

View full text Add to dashboard Cite

The transmembrane protein integrin plays a crucial role in the attachment of cells to the extracellular matrix. Integrin recruits many proteins intracellularly, including a four protein complex (Kindlin, Integrin linked kinase (ILK), PINCH, and parvin). C. elegans muscle provides an excellent model to study integrin adhesion complexes. In C. elegans, UNC-112 (Kindlin) binds to the cytoplasmic tail of PAT-3 (b-integrin) and to PAT-4 (ILK). We previously reported that PAT-4 binding to UNC-112 is essential for the binding of UNC-112 to PAT-3. Although there are crystal structures for ILK and a kindlin, there is no co-crystal structure available. To understand the molecular interaction between PAT-4 (ILK) and UNC-112 (Kindlin), we took a genetic approach. First, we isolated mutant PAT-4 proteins that cannot bind to UNC-112. Then, we isolated suppressor mutant UNC-112 proteins that restore interaction with mutant PAT-4 proteins. Second, these mutant PAT-4 proteins cannot localize to attachment structures in nematode muscle, but upon co-expression of an UNC-112 suppressor mutant protein, the mutant PAT-4 protein could localize to attachment structures. Third, overexpression of a mutant PAT-4 protein results in disorganization of the adhesion plaques at muscle cell boundaries, and co-expression of the UNC-112 supressor mutant protein alleviates this defect. Thus, we demonstrate that UNC-112 binding to PAT-4 is required for the localization and function of PAT-4 in integrin adhesion complexes in vivo. The missense mutations were mapped onto homology models of PAT-4 and UNC-112, and taking into account previously isolated mutations, we suggest a surface of PAT-4 that binds to UNC-112.

show abstract

Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 1 more Smart Citation

Genetic Analysis Suggests a Surface of PAT-4 (ILK) that Interacts with UNC-112 (kindlin)

Benian

Qadota

McPherson

et al. 2022

Preprint

Self Cite

View full text Add to dashboard Cite

show abstract

“…elegans are mostly transmitted laterally to the cuticle resulting in bending and sinuous locomotion of the worm on an agar surface [ 14 ]. In addition, integrin adhesion complexes exist at attachment plaques lying at the boundaries between adjacent muscle cells separated by basement membrane [ 15 ], and these also are likely to be involved in force transmission as their absence results in a locomotion defect [ 16 ].…”

Section: Introductionmentioning

confidence: 99%

Investigating the correlation of muscle function tests and sarcomere organization in C. elegans

et al. 2021

Self Cite

View full text Add to dashboard Cite

Background Caenorhabditis elegans has been widely used as a model to study muscle structure and function. Its body wall muscle is functionally and structurally similar to vertebrate skeletal muscle with conserved molecular pathways contributing to sarcomere structure, and muscle function. However, a systematic investigation of the relationship between muscle force and sarcomere organization is lacking. Here, we investigate the contribution of various sarcomere proteins and membrane attachment components to muscle structure and function to introduce C. elegans as a model organism to study the genetic basis of muscle strength. Methods We employ two recently developed assays that involve exertion of muscle forces to investigate the correlation of muscle function to sarcomere organization. We utilized a microfluidic pillar-based platform called NemaFlex that quantifies the maximum exertable force and a burrowing assay that challenges the animals to move in three dimensions under a chemical stimulus. We selected 20 mutants with known defects in various substructures of sarcomeres and compared the physiological function of muscle proteins required for force generation and transmission. We also characterized the degree of sarcomere disorganization using immunostaining approaches. Results We find that mutants with genetic defects in thin filaments, thick filaments, and M-lines are generally weaker, and our assays are successful in detecting the functional changes in response to each sarcomere location tested. We find that the NemaFlex and burrowing assays are functionally distinct informing on different aspects of muscle physiology. Specifically, the burrowing assay has a larger bandwidth in phenotyping muscle mutants, because it could pick ten additional mutants impaired while exerting normal muscle force in NemaFlex. This enabled us to combine their readouts to develop an integrated muscle function score that was found to correlate with the score for muscle structure disorganization. Conclusions Our results highlight the suitability of NemaFlex and burrowing assays for evaluating muscle physiology of C. elegans. Using these approaches, we discuss the importance of the studied sarcomere proteins for muscle function and structure. The scoring methodology we have developed enhances the utility of C. elegans as a genetic model to study muscle function.

show abstract

“…Unlike vertebrates, the contractile forces in C. elegans are mostly transmitted laterally to the cuticle resulting in bending and sinuous locomotion of the worm on an agar surface (14). In addition, integrin adhesion complexes exist at attachment plaques lying at the boundaries between adjacent muscle cells separated by basement membrane (15), and these also are likely to be involved in force transmission as their absence results in a locomotion defect (16).…”

Section: Introductionmentioning

confidence: 99%

Investigating the correlation of muscle function tests and sarcomere organization in C. elegans

Lesanpezeshki¹,

Qadota

Darabad³

et al. 2021

Preprint

Self Cite

View full text Add to dashboard Cite

Background: Caenorhabditis elegans has been widely used as a model to study muscle structure and function due to many genes having human homologs. Its body wall muscle is functionally and structurally similar to vertebrate skeletal muscle with conserved molecular pathways contributing to sarcomere structure, and muscle function. However, a systematic investigation of the relationship between muscle force and sarcomere organization is lacking. Here, we investigate the contribution of various sarcomere proteins and membrane attachment components to muscle structure and function to introduce C. elegans as a model organism to study the genetic basis of muscle strength. Methods: We employ two recently developed assays that involve exertion of muscle forces to investigate the correlation of muscle function to sarcomere organization. We utilized a microfluidic pillar-based platform called NemaFlex that quantifies the maximum exertable force and a burrowing assay that challenges the animals to move in three dimensions under a chemical stimulus. We selected 20 mutants with known defects in various substructures of sarcomeres and compared the physiological function of muscle proteins required for force generation and transmission. We also characterized the degree of sarcomere disorganization using immunostaining approaches. Results: We find that mutants with genetic defects in thin filaments, thick filaments and M-lines were generally weaker. Some mutations did not influence muscle strength suggesting a degree of redundancy in muscle genes contributing to muscle function. We find that the NemaFlex and burrowing assays are functionally distinct informing on different aspects of muscle physiology enabling us to combine their readouts to develop an integrated muscle function score. This muscle function score was found to negatively correlate with the score for muscle structure disorganization. Conclusions: Our results highlight the suitability of NemaFlex and burrowing assays for evaluating muscle physiology of C. elegans. Using these approaches, we discuss the importance of the studied sarcomere proteins for muscle function and structure. The scoring methodology we have developed lays the foundation for investigating the contribution of conserved sarcomere proteins and membrane attachment components to human muscle function and strength.

show abstract

The Rho-GEF PIX-1 directs assembly or stability of lateral attachment structures between muscle cells

Cited by 7 publications

References 53 publications

Genetic Analysis Suggests a Surface of PAT-4 (ILK) that Interacts with UNC-112 (kindlin)

Genetic Analysis Suggests a Surface of PAT-4 (ILK) that Interacts with UNC-112 (kindlin)

Investigating the correlation of muscle function tests and sarcomere organization in C. elegans

Investigating the correlation of muscle function tests and sarcomere organization in C. elegans

Contact Info

Product

Resources

About