The Tumor Suppressor CYLD Regulates Microtubule Dynamics and Plays a Role in Cell Migration

Abstract

The familial cylindromatosis tumor suppressor CYLD is known to contain three cytoskeleton-associated protein glycine-rich (CAP-Gly) domains, which exist in a number of microtubule-binding proteins and are responsible for their association with microtubules. However, it remains elusive whether CYLD interacts with microtubules and, if so, whether the interaction is mediated by the CAP-Gly domains. In this study, our data demonstrate that CYLD associates with microtubules both in cells and in vitro, and the first CAP-Gly domain of CYLD is mainly responsible for the interaction. Knockdown of cellular CYLD expression dramatically delays microtubule regrowth after nocodazole washout, indicating an activity for CYLD in promoting microtubule assembly. Our data further demonstrate that CYLD enhances tubulin polymerization into microtubules by lowering the critical concentration for microtubule assembly. In addition, we have identified by wound healing assay a critical role for CYLD in mediating cell migration and found that its first CAP-Gly domain is required for this activity. Thus CYLD joins a growing list of CAP-Gly domain-containing proteins that regulate microtubule dynamics and function.

CEP70 Protein Interacts with γ-Tubulin to Localize at the Centrosome and Is Critical for Mitotic Spindle Assembly

“…Nocodazole was then washed out, and microtubule regrowth was examined 1 or 5 min later by immunofluorescent staining with anti-␣-tubulin antibody and DAPI as described previously (13). For better visualization of microtubules, cells were treated with a buffer containing 100 mM PIPES, 1 mM EGTA, 0.5 mM MgCl 2 , and 0.5% Triton X-100 (pH 6.9) before immunofluorescent staining.…”

“…MBP-Cep70 was purified with the amylose resin according to the manufacturer's instructions (New England Biolabs). In vitro transcription and translation of ␥-tubulin were performed using the TNT SP6 high-yield wheat germ protein expression system (Promega) as described previously (13).…”

CYLD mediates ciliogenesis in multiple organs by deubiquitinating Cep70 and inactivating HDAC6

“…CYLD also contains cytoskeleton-associated protein glycine-rich (CAP-Gly) domains that mediate its interaction with microtubules and histone deacetylase 6 (HDAC6) [10][11][12]. Several cilium-related symptoms, such as male infertility, osteoporosis, and impaired lung maturation, have been reported in CYLD knockout mice or in mice carrying truncated CYLD [13][14][15].…”

CYLD coordinates with EB1 to regulate microtubule dynamics and cell migration

“…11,14 In addition, CYLD has been demonstrated to promote cell migration and thereby participate in normal physiological processes such as angiogenesis. 15 However, the molecular mechanisms linking the role of CYLD in the regulation of microtubule dynamics to its function in cell migration are largely unknown.…”

“…11 It consists of 956 amino acid residues with 3 cytoskeleton-associated protein glycine-rich (CAP-Gly) domains in the N terminus and a deubiquitinase domain in the C terminus. The canonical functions of CYLD Abbreviations: CYLD, cylindromatosis; EB1, end-binding protein 1; CLIP-170, cytoplasmic linker protein of 170 kDa; APC, adenomatous polyposis coli; +TIPs, microtubule plus end-tracking proteins; CAP-Gly, cytoskeleton-associated protein glycine-rich; IKBKG, inhibitor of kappa light polypeptide gene enhancer in B-cells, kinase gamma; TRAF, tumor necrosis factor receptorassociated factor; BCL3, B-cell chronic lymphocytic leukemia/lymphoma 3; NFκB, nuclear factor κB; HSPA1A, heat shock 70-kDa protein 1A; ABCB1, ATP-binding cassette sub-family B, member 1; STK11, serine/threonine kinase 11; siRNA, small interfering RNA; MAP, microtubule-associated protein; DAPI, 4',6-diamidino-2-phenylindole Cylindromatosis (CYLD), a deubiquitinase involved in inflammation and tumorigenesis via the modulation of cell signaling, has recently been identified as a critical regulator of microtubule dynamics.…”