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RASA1-driven cellular export of collagen IV is required for the development of lymphovenous and venous valves in mice.

ABSTRACT: RASA1, a negative regulator of Ras-MAPK signaling, is essential for the development and maintenance of lymphatic vessel valves. However, whether RASA1 is required for the development and maintenance of lymphovenous valves (LVV) and venous valves (VV) is unknown. In this study, we show that induced disruption of Rasa1 in mouse embryos did not affect initial specification of LVV or central VV, but did affect their continued development. Similarly, a switch to expression of a catalytically inactive form of RASA1 resulted in impaired LVV and VV development. Blocked development of LVV was associated with accumulation of the basement membrane protein, collagen IV, in LVV-forming endothelial cells (EC), and could be partially or completely rescued by MAPK inhibitors and drugs that promote collagen IV folding. Disruption of Rasa1 in adult mice resulted in venous hypertension and impaired VV function that was associated with loss of EC from VV leaflets. In conclusion, RASA1 functions as a negative regulator of Ras signaling in EC that is necessary for EC export of collagen IV, thus permitting the development of LVV and the development and maintenance of VV.

SUBMITTER: Chen D

PROVIDER: S-EPMC7746672 | biostudies-literature | 2020 Dec

REPOSITORIES: biostudies-literature

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RASA1-driven cellular export of collagen IV is required for the development of lymphovenous and venous valves in mice.

Chen Di D Geng Xin X Lapinski Philip E PE Davis Michael J MJ Srinivasan R Sathish RS King Philip D PD

Development (Cambridge, England) 20201207 23

RASA1, a negative regulator of Ras-MAPK signaling, is essential for the development and maintenance of lymphatic vessel valves. However, whether RASA1 is required for the development and maintenance of lymphovenous valves (LVV) and venous valves (VV) is unknown. In this study, we show that induced disruption of <i>Rasa1</i> in mouse embryos did not affect initial specification of LVV or central VV, but did affect their continued development. Similarly, a switch to expression of a catalytically i ...[more]

PMID: 33144395

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Project description:FlhA and FlhB are transmembrane proteins of the flagellar type III protein export apparatus, and their C-terminal cytoplasmic domains (FlhAC and FlhBC) coordinate flagellar protein export with assembly. FlhBC undergoes autocleavage between Asn-269 and Pro-270 in a well-conserved NPTH loop located between FlhBCN and FlhBCC polypeptides and interacts with the C-terminal domain of the FliK ruler when the length of the hook has reached about 55 nm in Salmonella As a result, the flagellar protein export apparatus switches its substrate specificity, thereby terminating hook assembly and initiating filament assembly. The mechanism of export switching remains unclear. Here, we report the role of FlhBC cleavage in the switching mechanism. Photo-cross-linking experiments revealed that the flhB(N269A) and flhB(P270A) mutations did not affect the binding affinity of FlhBC for FliK. Genetic analysis of the flhB(P270A) mutant revealed that the P270A mutation affects a FliK-dependent conformational change of FlhBC, thereby inhibiting the substrate specificity switching. The flhA(A489E) mutation in FlhAC suppressed the flhB(P270A) mutation, suggesting that an interaction between FlhBC and FlhAC is critical for the export switching. We propose that the interaction between FliKC and a cleaved form of FlhBC promotes a conformational change in FlhBC responsible for the termination of hook-type protein export and a structural remodeling of the FlhAC ring responsible for the initiation of filament-type protein export.IMPORTANCE The flagellar type III protein export apparatus coordinates protein export with assembly, which allows the flagellum to be efficiently built at the cell surface. Hook completion is an important morphological checkpoint for the sequential flagellar assembly process. The protein export apparatus switches its substrate specificity from the hook protein to the filament protein upon hook completion. FliK, FlhB, and FlhA are involved in the export-switching process, but the mechanism remains a mystery. By analyzing a slow-cleaving flhB(P270A) mutant, we provide evidence that an interaction between FliK and FlhB induces conformational rearrangements in FlhB, followed by a structural remodeling of the FlhA ring structure that terminates hook assembly and initiates filament formation.

Dataset Information

RASA1-driven cellular export of collagen IV is required for the development of lymphovenous and venous valves in mice.

Publications

RASA1-driven cellular export of collagen IV is required for the development of lymphovenous and venous valves in mice.

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