FAM83G/PAWS1 controls cytoskeletal dynamics and cell migration through association with the SH3 adaptor CD2AP

Sapkota, Gopal P.; Cummins, Timothy D.; Wu, Kevin Z. L.; Bozatzi, Polyxeni; Dingwell, Kevin S.; Macartney, Thomas; Wood, Nicola T.; Varghese, Joby; Gourlay, Robert; Campbell, David G.; Prescott, Alan R.; Griffis, Eric R.; Smith, James C.

doi:10.1101/106971

Cited by 5 publications

(6 citation statements)

References 35 publications

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“…1A; fig S1), we postulated that this domain might mediate the observed interaction between FAM83 and CK1 proteins. To map the minimal domain within FAM83 proteins that can interact with CK1 isoforms, we co-expressed Myc-tagged FAM83G fragments with full-length hemagglutinin (HA)-tagged CK1α in FAM83G -/- U2OS cells (28) and performed coimmunoprecipitation experiments (Fig. 3A).…”

Section: Resultsmentioning

confidence: 99%

“…The following constructs were generated: pcDNA5-FRT/TO GFP-FAM83A (DU44235), pcDNA5-FRT/TO GFP-FAM83B (DU44236), pcDNA5-FRT/TO GFP-FAM83C (DU42473), pcDNA5-FRT/TO GFP-FAM83D (DU42446), pcDNA5-FRT/TO GFP-FAM83E (DU44237), pcDNA5-FRT/TO GFP-FAM83F (DU44238), pcDNA5-FRT/TO GFP-FAM83G (DU33272), pcDNA5-FRT/TO GFP-FAM83H (DU44239), pcDNA5-FRT/TO GFP-FAM83C (D259A) (DU28479), pcDNA5-FRT/TO GFP-FAM83C (F293A) (DU28480), pcDNA5-FRT/TO GFP-FAM83E (D243A) (DU28481), pcDNA5-FRT/TO GFP-FAM83E (F277A) (DU28482), pcDNA5-FRT/TO GFP-FAM83F (D250A) (DU28268), pcDNA5-FRT/TO GFP-FAM83F (F284A) (DU28488), pcDNA5-FRT/TO GFP-FAM83G (D262A) (DU28476), pcDNA5-FRT/TO GFP-FAM83G (F296A) (DU28477), pcDNA5-FRT/TO GFP-FAM83H (D236A) (DU28428), pcDNA5-FRT/TO GFP-FAM83H (F270A) (DU28487), pcDNA5-FRT/TO mCherry-CK1α (DU28407), pcDNA5-FRT/TO mCherry-CK1α (N141A) (DU28839), pcDNA5-FRT/TO GFP-FAM83H(M1-L284)-FAM83G(S311-P823) (DU28683), pcDNA5-FRT/TO GFP-FAM83G(M1-V310)-FAM83H(V285-K1179) (DU28688), pcDNA5-FRT/TO FAM83A-GFP (DU42864), pcDNA5-FRT/TO FAM83B-GFP (DU42833), pcDNA5-FRT/TO FAM83C-GFP (DU42825), pcDNA5-FRT/TO FAM83D-GFP (DU42835), pcDNA5-FRT/TO FAM83E-GFP (DU42826), pcDNA5-FRT/TO FAM83F-GFP (DU42832), pcDNA5-FRT/TO FAM83G-GFP (DU42816), pcDNA5-FRT/TO FAM83H-GFP (DU42865), pcDNA5-FRT/TO GFP only (DU41455), pcDNA5-FRT/TO GFP-FAM83H (F274A) (DU28658), pcDNA5-FRT/TO GFP-FAM83H (F270, 274A) (DU28182), pcDNA5-FRT/TO FLAG-FAM83G (DU33274), pcDNA5-FRT/TO FLAG-FAM83G (F296A) (DU28024), pcDNA5-FRT/TO FLAG-FAM83G (F296A, F300A) (DU28026), pcDNA5-FRT/TO FLAG-FAM83G (F300A) (DU28025), pCS2+ HA-CK1α (DU28216), pCMV5-FLAG TTBK2 (DU19028), pCMV-FLAG-CK1γ (DU5580), pCS2+ HA CK1δ (DU28189), pcDNA5-FRT/TO mcherry-CK1ε (DU28406). Myc-xFAM83G (Xenopus laevis FAM83G) constructs have been described previously (28). For CRISPR/Cas9 gene editing, pBABED P U6 FAM83H KO sense gRNA (DU52010), pX335-CAS9-D10A FAM83H KO antisense gRNA (DU52026), pBABED P U6 FAM83G KI sense gRNA (DU48528), pX335-Cas9-D10A FAM83G KI antisense gRNA (DU48529), pEX-K4 FAM83G Cter GFP donor (DU48585), pBABED P U6 FAM83B KI sense gRNA (DU54494), pX335-Cas9-D10A FAM83B KI antisense gRNA (DU54504), and pEX-K4 FAM83B Nter GFP donor (DU54547) were generated.…”

Section: Methodsmentioning

confidence: 99%

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The DUF1669 domain of FAM83 family proteins anchor casein kinase 1 isoforms

et al. 2018

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Members of the casein kinase 1 (CK1) family of serine-threonine protein kinases are implicated in the regulation of many cellular processes, including the cell cycle, circadian rhythms, and Wnt and Hedgehog signaling. Because these kinases exhibit constitutive activity in biochemical assays, it is likely that their activity in cells is controlled by subcellular localization, interactions with inhibitory proteins, targeted degradation, or combinations of these mechanisms. We identified members of the FAM83 family of proteins as partners of CK1 in cells. All eight members of the FAM83 family (FAM83A to FAM83H) interacted with the α and α-like isoforms of CK1; FAM83A, FAM83B, FAM83E, and FAM83H also interacted with the δ and ε isoforms of CK1. We detected no interaction between any FAM83 member and the related CK1γ1, CK1γ2, and CK1γ3 isoforms. Each FAM83 protein exhibited a distinct pattern of subcellular distribution and colocalized with the CK1 isoform(s) to which it bound. The interaction of FAM83 proteins with CK1 isoforms was mediated by the conserved domain of unknown function 1669 (DUF1669) that characterizes the FAM83 family. Mutations in FAM83 proteins that prevented them from binding to CK1 interfered with the proper subcellular localization and cellular functions of both the FAM83 proteins and their CK1 binding partners. On the basis of its function, we propose that DUF1669 be renamed the polypeptide anchor of CK1 domain.

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Section: Resultsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

The DUF1669 domain of FAM83 family proteins anchor casein kinase 1 isoforms

et al. 2018

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“…PAWS1 deficiency causes severe defects in F-actin organisation and distribution, as well as lamellipodial organisation, resulting in impaired cell migration. The dynamic association of PAWS1 with the actin/cytoskeletal regulator CD2AP at lamellae appears to be essential for PAWS1 to control actin organisation and cellular migration [ 36 ]. Interestingly, deficiency of CD2AP phenocopies the actin and cell migration defects observed when cells are depleted of PAWS1 [ 36 ].…”

Section: The Role Of Fam83g In Cell Signalling Morphology and Diseasmentioning

confidence: 99%

“…The dynamic association of PAWS1 with the actin/cytoskeletal regulator CD2AP at lamellae appears to be essential for PAWS1 to control actin organisation and cellular migration [ 36 ]. Interestingly, deficiency of CD2AP phenocopies the actin and cell migration defects observed when cells are depleted of PAWS1 [ 36 ]. Recently, we showed that PAWS1 overexpression in Xenopus embryos activates Wnt signalling and causes complete axis duplication.…”

Section: The Role Of Fam83g In Cell Signalling Morphology and Diseasmentioning

confidence: 99%

The FAM83 family of proteins: from pseudo-PLDs to anchors for CK1 isoforms

Bozatzi

Sapkota

2018

Biochemical Society Transactions

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The eight members of the FAM83 (FAMily with sequence similarity 83) family of poorly characterised proteins are only present in vertebrates and are defined by the presence of the conserved DUF1669 domain of unknown function at their N-termini. The DUF1669 domain consists of a conserved phospholipase D (PLD)-like catalytic motif. However, the FAM83 proteins display no PLD catalytic (PLDc) activity, and the pseudo-PLDc motif present in each FAM83 member lacks the crucial elements of the native PLDc motif. In the absence of catalytic activity, it is likely that the DUF1669 domain has evolved to espouse novel function(s) in biology. Recent studies have indicated that the DUF1669 domain mediates the interaction with different isoforms of the CK1 (casein kinase 1) family of Ser/Thr protein kinases. In turn, different FAM83 proteins, which exhibit unique amino acid sequences outside the DUF1669 domain, deliver CK1 isoforms to unique subcellular compartments. One of the first protein kinases to be discovered, the CK1 isoforms are thought to be constitutively active and are known to control a plethora of biological processes. Yet, their regulation of kinase activity, substrate selectivity and subcellular localisation has remained a mystery. The emerging evidence now supports a central role for the DUF1669 domain, and the FAM83 proteins, in the regulation of CK1 biology.

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“…CD2-associated protein (CD2AP) is a cortactin-binding scaffold protein that controls actin dynamics and participates in membrane trafficking during endocytosis by interacting with dynamin, synaptojanin1, endophilin, and nephrin ( 49 , 95 ). CD2AP has three Src homology 3 (SH3) domains and a coiled-coil domain contributing to its functions on actin dynamics and endocytosis by interactions with other proteins containing proline-rich domains ( 96 ). Interestingly, mice lacking Cd2ap develop nephrotic syndrome and die at 6–7 weeks of age due to kidney failure ( 21 ).…”

Section: Endocytosis In Podocyte Homeostasis and Diseasementioning

confidence: 99%

Podocyte Endocytosis in Regulating the Glomerular Filtration Barrier

2022

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Endocytosis is a mechanism that internalizes and recycles plasma membrane components and transmembrane receptors via vesicle formation, which is mediated by clathrin-dependent and clathrin-independent signaling pathways. Podocytes are specialized, terminally differentiated epithelial cells in the kidney, located on the outermost layer of the glomerulus. These cells play an important role in maintaining the integrity of the glomerular filtration barrier in conjunction with the adjacent basement membrane and endothelial cell layers within the glomerulus. An intact podocyte endocytic machinery appears to be necessary for maintaining podocyte function. De novo pathologic human genetic mutations and loss-of-function studies of critical podocyte endocytosis genes in genetically engineered mouse models suggest that this pathway contributes to the pathophysiology of development and progression of proteinuria in chronic kidney disease. Here, we review the mechanism of cellular endocytosis and its regulation in podocyte injury in the context of glomerular diseases. A thorough understanding of podocyte endocytosis may shed novel insights into its biological function in maintaining a functioning filter and offer potential targeted therapeutic strategies for proteinuric glomerular diseases.

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FAM83G/PAWS1 controls cytoskeletal dynamics and cell migration through association with the SH3 adaptor CD2AP

Cited by 5 publications

References 35 publications

The DUF1669 domain of FAM83 family proteins anchor casein kinase 1 isoforms

The DUF1669 domain of FAM83 family proteins anchor casein kinase 1 isoforms

The FAM83 family of proteins: from pseudo-PLDs to anchors for CK1 isoforms

Podocyte Endocytosis in Regulating the Glomerular Filtration Barrier

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