{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,10,3]],"date-time":"2026-10-03T07:42:23Z","timestamp":1791013343256,"version":"4.1.0"},"reference-count":35,"publisher":"Wiley","issue":"1","license":[{"start":{"date-parts":[[2004,2,19]],"date-time":"2004-02-19T00:00:00Z","timestamp":1077148800000},"content-version":"vor","delay-in-days":3336,"URL":"http:\/\/onlinelibrary.wiley.com\/termsAndConditions#vor"}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["J of Cellular Biochemistry"],"published-print":{"date-parts":[[1995,1]]},"abstract":"<jats:title>Abstract<\/jats:title><jats:p>SPARC (secreted protein, acidic and rich in cysteine, also known as osteonectin and BM\u201040) is a metal\u2010binding glycoprotein secreted by a variety of cultured cells and characteristic of tissues undergoing morphogenesis, remodeling, and repair. Recently it has been shown that SPARC inhibits the progression of the endothelial cell cycle in mid\u2010G<jats:sub>1<\/jats:sub>, and that a synthetic peptide (amino acids 54\u201373 of secreted murine SPARC, peptide 2.1) from a cationic, disulfide\u2010bonded region was in part responsible for the growth\u2010suppressing activity [Funk and Sage (1991): Proc Natl Acad Sci USA 88:2648\u20132652]. Moreover, SPARC was shown to interact directly with bovine aortic endothelial (BAE) cells through a C\u2010terminal EF\u2010hand sequence comprising a high\u2010affinity Ca<jats:sup>2+<\/jats:sup>\u2010binding site of SPARC and represented by a synthetic peptide (amino acids 254\u2013273) termed 4.2 [Yost and Sage (1993): J Biol Chem 268:25790\u201325796]. In this study we show that peptide 4.2 is a more potent inhibitor of DNA synthesis that acts cooperatively with peptide 2.1 to diminish the incorporation of [<jats:sup>3<\/jats:sup>H]\u2010thymidine by both BAE and bovine capillary endothelial (BCE) cells. At concentrations of 0.019\u20130.26 mM peptide 4.2, thymidine incorporation by BAE cells was decreased incrementally, relative to control values, from approximately 100 to 10%. Although somewhat less responsive, BCE cells exhibited a dose\u2010responsive decrement in thymidine incorporation, with a maximal inhibition of 55% at 0.39 mM. The inhibitory effect of peptide 4.2 was essentially independent of heparin and basic fibroblast growth factor and was blocked by anti\u2010SPARC peptide 4.2 IgG, but not by antibodies specific for other domains of SPARC. To identify residues that were necessary for inhibition of DNA synthesis, we introduced single amino acid substitutions into synthetic peptide 4.2 and tested their activities and cell\u2010surface binding characteristics on endothelial cells. Two peptides displayed null to diminished effects in the bioassays that were concentration\u2010dependent: peptide 4.2 K, containing an Asp<jats:sub>258<\/jats:sub> \u2192 Lys substitution, and peptide 4.2 AA, in which the two disulfide\u2010bonded Cys (positions 255 and 271) were changed to Ala residues. Peptide 4.2 K, which failed to fulfill the EF\u2010hand consensus formula, exhibited an anomalous fluorescence emission spectrum, in comparison with the wild\u2010type 4.2 sequence, that was indicative of a compromised affinity for Ca<jats:sup>2+<\/jats:sup>. Moreover, ablation of the disulfide bond in peptide 4.2 AA potentially destabilized the Ca<jats:sup>2+<\/jats:sup>\u2010binding loop structure, as assessed by fluorescence spectroscopy, such that the peptide competed poorly for the binding of [<jats:sup>125<\/jats:sup>I]\u2010peptide 4.2 to BAE cells. We conclude both that Ca<jats:sup>2+<\/jats:sup>\u2010coordinating Asp at position 258 and the conformation of peptide 4.2 are necessary for the inhibition of DNA synthesis by SPARC in cultured endothelial cells.<\/jats:p>","DOI":"10.1002\/jcb.240570113","type":"journal-article","created":{"date-parts":[[2005,5,28]],"date-time":"2005-05-28T21:59:37Z","timestamp":1117317577000},"page":"127-140","source":"Crossref","is-referenced-by-count":55,"title":["Inhibition of endothelial cell proliferation by SPARC is mediated through a Ca<sup>2+<\/sup>\u2010binding EF\u2010hand sequence"],"prefix":"10.1002","volume":"57","author":[{"given":"E. Helene","family":"Sage","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"James A.","family":"Bassuk","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Jeffrey C.","family":"Yost","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"M. Judah","family":"Folkman","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Timothy F.","family":"Lane","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"311","published-online":{"date-parts":[[2004,2,19]]},"reference":[{"key":"e_1_2_1_2_1","doi-asserted-by":"publisher","DOI":"10.1093\/nar\/20.suppl.2013"},{"key":"e_1_2_1_3_1","doi-asserted-by":"publisher","DOI":"10.1016\/0955-0674(91)90053-2"},{"key":"e_1_2_1_4_1","doi-asserted-by":"crossref","first-page":"24067","DOI":"10.1016\/S0021-9258(20)80493-1","article-title":"Differential recovery of Ca2+ binding activity in mutated EF\u2010hands of cardiac toponin C","volume":"268","author":"Dotson DG","year":"1993","journal-title":"J Biol Chem"},{"key":"e_1_2_1_5_1","doi-asserted-by":"publisher","DOI":"10.1021\/bi00396a015"},{"key":"e_1_2_1_6_1","doi-asserted-by":"publisher","DOI":"10.1091\/mbc.3.10.1057"},{"key":"e_1_2_1_7_1","doi-asserted-by":"publisher","DOI":"10.1016\/S0021-9258(19)49853-0"},{"key":"e_1_2_1_8_1","doi-asserted-by":"publisher","DOI":"10.1073\/pnas.88.7.2648"},{"key":"e_1_2_1_9_1","doi-asserted-by":"publisher","DOI":"10.1002\/jcp.1041540108"},{"key":"e_1_2_1_10_1","doi-asserted-by":"publisher","DOI":"10.1073\/pnas.91.8.3448"},{"key":"e_1_2_1_11_1","doi-asserted-by":"publisher","DOI":"10.1038\/345132a0"},{"key":"e_1_2_1_12_1","doi-asserted-by":"crossref","first-page":"13178","DOI":"10.1016\/S0021-9258(18)98821-6","article-title":"SPARC induces the expression of type I plasminogen activator inhibitor in cultured bovine aortic endothelial cells","volume":"266","author":"Hasselaar P","year":"1991","journal-title":"J Biol Chem"},{"key":"e_1_2_1_13_1","doi-asserted-by":"publisher","DOI":"10.1002\/jcb.240490310"},{"key":"e_1_2_1_14_1","first-page":"174","article-title":"Differential expression of extracellular proteins is correlated with angiogenesis in vitro","volume":"64","author":"Iruela\u2010Arispe M\u2010L","year":"1991","journal-title":"Lab Invest"},{"key":"e_1_2_1_15_1","doi-asserted-by":"publisher","DOI":"10.1083\/jcb.111.6.3065"},{"key":"e_1_2_1_16_1","doi-asserted-by":"crossref","first-page":"16736","DOI":"10.1016\/S0021-9258(18)42063-7","article-title":"Regulation of gene expression by SPARC during angiogenesis in vitro: Fibronectin, thrombospondin, and type 1 plasminogen activator inhibitor","volume":"267","author":"Lane TF","year":"1992","journal-title":"J Biol Chem"},{"key":"e_1_2_1_17_1","doi-asserted-by":"publisher","DOI":"10.1096\/fasebj.8.2.8119487"},{"key":"e_1_2_1_18_1","doi-asserted-by":"publisher","DOI":"10.1083\/jcb.125.4.929"},{"key":"e_1_2_1_19_1","doi-asserted-by":"publisher","DOI":"10.1139\/o90-084"},{"key":"e_1_2_1_20_1","doi-asserted-by":"publisher","DOI":"10.1021\/bi00149a006"},{"key":"e_1_2_1_21_1","doi-asserted-by":"publisher","DOI":"10.1111\/j.1432-1033.1992.tb16773.x"},{"key":"e_1_2_1_22_1","doi-asserted-by":"publisher","DOI":"10.1111\/j.1432-1033.1991.tb15996.x"},{"key":"e_1_2_1_23_1","doi-asserted-by":"publisher","DOI":"10.1083\/jcb.115.4.1127"},{"key":"e_1_2_1_24_1","doi-asserted-by":"publisher","DOI":"10.1002\/jez.1402650207"},{"key":"e_1_2_1_25_1","doi-asserted-by":"publisher","DOI":"10.1073\/pnas.89.4.1281"},{"key":"e_1_2_1_26_1","doi-asserted-by":"crossref","first-page":"14831","DOI":"10.1016\/S0021-9258(18)98545-5","article-title":"Minireview: Extracellular proteins that modulate cell\u2010matrix interactions: SPARC, tenascin, and thrombospondin","volume":"266","author":"Sage EH","year":"1991","journal-title":"J Biol Chem"},{"key":"e_1_2_1_27_1","doi-asserted-by":"publisher","DOI":"10.1016\/S0021-9258(17)43194-2"},{"key":"e_1_2_1_28_1","doi-asserted-by":"publisher","DOI":"10.1177\/37.6.2723400"},{"key":"e_1_2_1_29_1","doi-asserted-by":"publisher","DOI":"10.1083\/jcb.109.1.341"},{"key":"e_1_2_1_30_1","doi-asserted-by":"publisher","DOI":"10.1091\/mbc.4.9.941"},{"key":"e_1_2_1_31_1","doi-asserted-by":"publisher","DOI":"10.1016\/0076-6879(91)98011-T"},{"key":"e_1_2_1_32_1","doi-asserted-by":"publisher","DOI":"10.1126\/science.6199844"},{"key":"e_1_2_1_33_1","doi-asserted-by":"publisher","DOI":"10.1021\/bi00064a008"},{"key":"e_1_2_1_34_1","doi-asserted-by":"publisher","DOI":"10.1083\/jcb.121.6.1433"},{"key":"e_1_2_1_35_1","doi-asserted-by":"publisher","DOI":"10.1002\/jbmr.5650040612"},{"key":"e_1_2_1_36_1","doi-asserted-by":"crossref","first-page":"25790","DOI":"10.1016\/S0021-9258(19)74459-7","article-title":"Specific interaction of SPARC with endothelial cells is mediated through a carboxy\u2010terminal sequence containing a calcium\u2010binding E\u2010F hand","volume":"268","author":"Yost JC","year":"1993","journal-title":"J Biol Chem"}],"container-title":["Journal of Cellular Biochemistry"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/api.wiley.com\/onlinelibrary\/tdm\/v1\/articles\/10.1002%2Fjcb.240570113","content-type":"unspecified","content-version":"vor","intended-application":"text-mining"},{"URL":"https:\/\/onlinelibrary.wiley.com\/doi\/pdf\/10.1002\/jcb.240570113","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2023,10,25]],"date-time":"2023-10-25T17:25:48Z","timestamp":1698254748000},"score":1,"resource":{"primary":{"URL":"https:\/\/onlinelibrary.wiley.com\/doi\/10.1002\/jcb.240570113"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[1995,1]]},"references-count":35,"journal-issue":{"issue":"1","published-print":{"date-parts":[[1995,1]]}},"alternative-id":["10.1002\/jcb.240570113"],"URL":"https:\/\/doi.org\/10.1002\/jcb.240570113","archive":["Portico"],"relation":{},"ISSN":["0730-2312","1097-4644"],"issn-type":[{"value":"0730-2312","type":"print"},{"value":"1097-4644","type":"electronic"}],"subject":[],"published":{"date-parts":[[1995,1]]}}}