Abstract
Podosome-type adhesions are actin-based membrane protrusions involved in cell-matrix adhesion and extracellular matrix degradation. Despite growing knowledge of many proteins associated with podosome-type adhesions, much remains unknown concerning the function of podosomal proteins at the level of the whole animal. In this study, the spontaneous mouse mutant nee was used to identify a component of podosome-type adhesions that is essential for normal postnatal growth and development. Mice homozygous for the nee allele exhibited runted growth, craniofacial and skeletal abnormalities, ocular anterior segment dysgenesis, and hearing impairment. Adults also exhibited infertility and a form of lipodystrophy. Using genetic mapping and DNA sequencing, the cause of nee phenotypes was identified as a 1-bp deletion within the Sh3pxd2b gene on mouse Chromosome 11. Whereas the wild-type Sh3pxd2b gene is predicted to encode a protein with one PX domain and four SH3 domains, the nee mutation is predicted to cause a frameshift and a protein truncation altering a portion of the third SH3 domain and deleting all of the fourth SH3 domain. The SH3PXD2B protein is believed to be an important component of podosomes likely to mediate protein-protein interactions with membrane-spanning metalloproteinases. Testing this directly, SH3PXD2B localized to podosomes in constitutively active Src-transfected fibroblasts and through its last SH3 domain associated with a transmembrane member of a disintegrin and metalloproteinase family of proteins, ADAM15. These results identify SH3PXD2B as a podosomal-adaptor protein required for postnatal growth and development, particularly within physiologic contexts involving extracellular matrix regulation.
This is a preview of subscription content, log in via an institution to check access.
Similar content being viewed by others
References
Abram CL, Seals DF, Pass I, Salinsky D, Maurer L et al (2003) The adaptor protein fish associates with members of the ADAMs family and localizes to podosomes of Src-transformed cells. J Biol Chem 278:16844–16851
Acott TS, Kelley MJ (2008) Extracellular matrix in the trabecular meshwork. Exp Eye Res 86:543–561
Aga M, Bradley JM, Keller K, Kelley MJ, Acott TS (2008) Specialized podosome- or invadopodia-like structures (PILS) for focal trabecular meshwork extracellular matrix turnover. Invest Ophthalmol Vis Sci 49:5353–5365
Bourlier V, Zakaroff-Girard A, De Barros S, Pizzacalla C, de Saint Front VD et al (2005) Protease inhibitor treatments reveal specific involvement of matrix metalloproteinase-9 in human adipocyte differentiation. J Pharmacol Exp Ther 312:1272–1279
Buffone GJ, Darlington GJ (1985) Isolation of DNA from biological specimens without extraction with phenol. Clin Chem 31:164–165
Buschman MD, Bromann PA, Cejudo-Martin P, Wen F, Pass I et al (2009) The novel adaptor protein Tks4 (SH3PXD2B) is required for functional podosome formation. Mol Biol Cell 20:1302–1311
Chang B, Hawes NL, Hurd RE, Wang J, Howell D et al (2005) Mouse models of ocular diseases. Vis Neurosci 22:587–593
Chen W, Yang S, Abe Y, Li M, Wang Y et al (2007) Novel pycnodysostosis mouse model uncovers cathepsin K function as a potential regulator of osteoclast apoptosis and senescence. Hum Mol Genet 16:410–423
Chun TH, Hotary KB, Sabeh F, Saltiel AR, Allen ED et al (2006) A pericellular collagenase directs the 3-dimensional development of white adipose tissue. Cell 125:577–591
Courtneidge SA, Azucena EF, Pass I, Seals DF, Tesfay L (2005) The SRC substrate Tks5, podosomes (invadopodia), and cancer cell invasion. Cold Spring Harb Symp Quant Biol 70:167–171
Destaing O, Sanjay A, Itzstein C, Horne WC, Toomre D et al (2008) The tyrosine kinase activity of c-Src regulates actin dynamics and organization of podosomes in osteoclasts. Mol Biol Cell 19:394–404
Egeblad M, Shen HC, Behonick DJ, Wilmes L, Eichten A, Korets LV, Kheradmand F, Werb Z, Coussens LM (2007) Type I collagen is a genetic modifier of matrix metalloproteinase 2 in murine skeletal development. Dev Dyn 236:1683–1693
Eto K, Huet C, Tarui T, Kupriyanov S, Liu HZ et al (2002) Functional classification of ADAMs based on a conserved motif for binding to integrin alpha 9beta 1: implications for sperm-egg binding and other cell interactions. J Biol Chem 277:17804–17810
Evans JP (2001) Fertilin beta and other ADAMs as integrin ligands: insights into cell adhesion and fertilization. Bioessays 23:628–639
Gimona M, Buccione R (2006) Adhesions that mediate invasion. Int J Biochem Cell Biol 38:1875–1892
Gimona M, Buccione R, Courtneidge SA, Linder S (2008) Assembly and biological role of podosomes and invadopodia. Curr Opin Cell Biol 20:235–241
Green M (1952) A rapid method for clearing and staining specimens for the demonstration of bone. Ohio J Sci 52:31–33
Grisendi S, Bernardi R, Rossi M, Cheng K, Khandker L et al (2005) Role of nucleophosmin in embryonic development and tumorigenesis. Nature 437:147–153
Hikita A, Yana I, Wakeyama H, Nakamura M, Kadono Y et al (2006) Negative regulation of osteoclastogenesis by ectodomain shedding of receptor activator of NF-kappaB ligand. J Biol Chem 281:36846–36855
Hishida T, Eguchi T, Osada S, Nishizuka M, Imagawa M (2008) A novel gene, fad49, plays a crucial role in the immediate early stage of adipocyte differentiation via involvement in mitotic clonal expansion. FEBS J 275:5576–5588
Holmbeck K, Bianco P, Caterina J, Yamada S, Kromer M et al (1999) MT1-MMP-deficient mice develop dwarfism, osteopenia, arthritis, and connective tissue disease due to inadequate collagen turnover. Cell 99:81–92
Kornak U, Kasper D, Bosl MR, Kaiser E, Schweizer M et al (2001) Loss of the ClC-7 chloride channel leads to osteopetrosis in mice and man. Cell 104:205–215
Krane SM, Inada M (2008) Matrix metalloproteinases and bone. Bone 43:7–18
Linder S (2007) The matrix corroded: podosomes and invadopodia in extracellular matrix degradation. Trends Cell Biol 17:107–117
Linder S, Aepfelbacher M (2003) Podosomes: adhesion hot-spots of invasive cells. Trends Cell Biol 13:376–385
Liu YJ, Liu XG, Wang L, Dina C, Yan H et al (2008) Genome-wide association scans identified CTNNBL1 as a novel gene for obesity. Hum Mol Genet 17:1803–1813
Lock P, Abram CL, Gibson T, Courtneidge SA (1998) A new method for isolating tyrosine kinase substrates used to identify fish, an SH3 and PX domain-containing protein, and Src substrate. EMBO J 17:4346–4357
Malinin NL, Wright S, Seubert P, Schenk D, Griswold-Prenner I (2005) Amyloid-beta neurotoxicity is mediated by FISH adapter protein and ADAM12 metalloprotease activity. Proc Natl Acad Sci USA 102:3058–3063
Mosig RA, Dowling O, DiFeo A, Ramirez MC, Parker IC, Abe E, Diouri J, Aqeel AA, Wylie JD, Oblander SA, Madri J, Bianco P, Apte SS, Zaidi M, Doty SB, Majeska RJ, Schafler MB, Martignetti JA (2007) Loss of MMP-2 disrupts skeletal and craniofacial development and results in decreased bone mineralization, joint erosion and defects in osteoblast and osteoclast growth. Hum Mol Genet 16:1113–1123
Ohbayashi N, Shibayama M, Kurotaki Y, Imanishi M, Fujimori T et al (2002) FGF18 is required for normal cell proliferation and differentiation during osteogenesis and chondrogenesis. Genes Dev 16:870–879
Oikawa T, Itoh T, Takenawa T (2008) Sequential signals toward podosome formation in NIH-src cells. J Cell Biol 182:157–169
Ory S, Brazier H, Pawlak G, Blangy A (2008) Rho GTPases in osteoclasts: orchestrators of podosome arrangement. Eur J Cell Biol 87:469–477
Pasten-Hidalgo K, Hernandez-Rivas R, Roa-Espitia AL, Sanchez-Gutierrez M, Martinez-Perez F et al (2008) Presence, processing, and localization of mouse ADAM15 during sperm maturation and the role of its disintegrin domain during sperm-egg binding. Reproduction 136:41–51
Richtsmeier JT, Baxter LL, Reeves RH (2000) Parallels of craniofacial maldevelopment in Down syndrome and Ts65Dn mice. Dev Dyn 217:137–145
Rocha JL, Eisen EJ, Van Vleck LD, Pomp D (2004) A large-sample QTL study in mice: I. Growth. Mamm Genome 15:83–99
Saltel F, Chabadel A, Bonnelye E, Jurdic P (2008) Actin cytoskeletal organisation in osteoclasts: a model to decipher transmigration and matrix degradation. Eur J Cell Biol 87:459–468
Sharma VM, Litersky JM, Bhaskar K, Lee G (2007) Tau impacts on growth-factor-stimulated actin remodeling. J Cell Sci 120:748–757
Shi J, Son MY, Yamada S, Szabova L, Kahan S et al (2008) Membrane-type MMPs enable extracellular matrix permissiveness and mesenchymal cell proliferation during embryogenesis. Dev Biol 313:196–209
Shimizu A, Maruyama T, Tamaki K, Uchida H, Asada H et al (2005) Impairment of decidualization in SRC-deficient mice. Biol Reprod 73:1219–1227
Soriano P, Montgomery C, Geske R, Bradley A (1991) Targeted disruption of the c-src proto-oncogene leads to osteopetrosis in mice. Cell 64:693–702
Spinardi L, Marchisio PC (2006) Podosomes as smart regulators of cellular adhesion. Eur J Cell Biol 85:191–194
Symons M (2008) Cell biology: watching the first steps of podosome formation. Curr Biol 18:R925–R927
Tarone G, Cirillo D, Giancotti FG, Comoglio PM, Marchisio PC (1985) Rous sarcoma virus-transformed fibroblasts adhere primarily at discrete protrusions of the ventral membrane called podosomes. Exp Cell Res 159:141–157
Thompson O, Kleino I, Crimaldi L, Gimona M, Saksela K et al (2008) Dystroglycan, Tks5 and Src mediated assembly of podosomes in myoblasts. PLoS ONE 3:e3638
Vogel CI, Greene B, Scherag A, Muller TD, Friedel S et al (2009) Non-replication of an association of CTNNBL1 polymorphisms and obesity in a population of Central European ancestry. BMC Med Genet 10:14
Weaver AM (2008) Invadopodia. Curr Biol 18:R362–R364
Young JS, Guttman JA, Vaid KS, Shahinian H, Vogl AW (2009a) Cortactin (CTTN), N-WASP (WASL), and clathrin (CLTC) are present at podosome-like tubulobulbar complexes in the rat testis. Biol Reprod 80:153–161
Young JS, Guttman JA, Vaid KS, Vogl AW (2009b) Tubulobulbar complexes are intercellular podosome-like structures that internalize intact intercellular junctions during epithelial remodeling events in the rat testis. Biol Reprod 80:162–174
Zhao LJ, Xiao P, Liu YJ, Xiong DH, Shen H et al (2007) A genome-wide linkage scan for quantitative trait loci underlying obesity related phenotypes in 434 Caucasian families. Hum Genet 121:145–148
Zheng QY, Johnson KR, Erway LC (1999) Assessment of hearing in 80 inbred strains of mice by ABR threshold analyses. Hear Res 130:94–107
Zheng QY, Hardisty-Hughes R, Brown SD (2006) Mouse models as a tool to unravel the genetic basis for human otitis media. Brain Res 1091:9–15
Acknowledgments
We thank Adam Hedberg-Buenz for maintaining mouse colonies at the University of Iowa; Pat Ward-Bailey for help with the nee mapping cross; John Fingert and Val Sheffield for contributions to DNA sequencing; Gloria Lee for kindly providing the Src (Y529F) construct; Michael Henry for assistance with X-ray imaging; Wayne Johnson and Robert Mullins for helpful discussions; and Michelle Curtain, an animal caretaker at The Jackson Laboratory, who observed the founder nee mouse and provided it to The Jackson Laboratory’s Mouse Mutant Resource. We are especially grateful for many helpful discussions with Norm Hawes of The Jackson Laboratory, who recently passed away; his selfless attitude toward science and kindness were a joy and inspiration. This work was supported by EY017673 and a grant from the Knights Templar Eye Foundation, Inc. to MGA. The Mouse Mutant Resource of The Jackson Laboratory is supported by a grant from the National Center for Research Resources (RR01183); contributions to work on the craniofacial and skeletal phenotypes by LRD and BH were supported by EYO15073; contributions to the assessment of the hearing phenotype by QYZ and KRJ were supported by DC005846 and DC004301; contributions to initially recognizing ocular phenotypes of nee mice by BC were supported by the Foundation Fighting Blindness; and work at The Jackson Laboratory was additionally supported by National Cancer Institute Grant CA34196.
Author information
Authors and Affiliations
Corresponding author
Rights and permissions
About this article
Cite this article
Mao, M., Thedens, D.R., Chang, B. et al. The podosomal-adaptor protein SH3PXD2B is essential for normal postnatal development. Mamm Genome 20, 462–475 (2009). https://doi.org/10.1007/s00335-009-9210-9
Received:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s00335-009-9210-9