Developmentally regulated protease expression during sea urchin embryogenesis.
Developmentally regulated protease expression during sea urchin embryogenesis.
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海胆胚胎发生过程中发育调节的蛋白酶表达。
DOI：
10.1002/mrd.1080400106
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发表时间：
1995
期刊：
影响因子：
--
通讯作者：
Nishioka,D
中科院分区：
文献类型：
--
作者：
Vafa,O;Nishioka,D
采用SDS-PAGE明胶底物酶谱技术对蛋白酶表达进行的时间研究显示，在海胆，Lytechinus pictus和Strongylocentrotus purpuratus的早期发育过程中，蛋白酶出现了明确的外观。这些蛋白酶的水平在每个物种的原肠胚形成过程中大幅增加。发现相对分子质量为57和50 kDa的两种主要蛋白酶被锌螯合剂1，10-菲咯啉、更非特异性的金属螯合剂EDTA和还原剂二硫苏糖醇抑制。丝氨酸蛋白酶抑制剂，苯甲脒，这些蛋白酶的活性没有产生影响，这两种酶表现出活性在中性到弱碱性pH范围。用放线菌素D（一种转录抑制剂）处理胚胎，受精后9小时开始，受精后48小时抑制了随后出现的两种蛋白酶，以及与原肠胚形成相关的任何形态学变化。受精后15和21小时开始的治疗导致蛋白酶水平的增加，这些蛋白酶与在原肠形成的连续更高级阶段的逮捕相关。5种不同胚胎组分的SDS-PAGE酶谱分析表明，57-和50-kDa蛋白酶位于囊胚腔中，囊胚腔蛋白酶活性通过原位酶谱进一步在显微镜下证实。因此，57-和50-kDa蛋白酶被表征为金属蛋白酶。它们的表达取决于胚胎基因组的转录，它们的时空出现表明它们参与原肠胚形成期间的囊胚腔基质重塑。© 1995 Wiley利斯公司
A temporal study of protease expression employing the technique of SDS‐PAGE gelatin substrate zymography revealed a definitive appearance of proteases during early development in the sea urchins,Lytechinus pictusandStrongylocentrotus purpuratus. The levels of these proteases increase substantially during gastrulation in each species. The two major proteases with relative molecular masses of 57 and 50 kDa were found to be inhibited by the zinc chelator, 1,10‐phenanthroline, the more nonspecific metal chelator, EDTA, and the reducing agent, dithiothreitol. The serine protease inhibitor, benzamidine, exerted no effects on the activities of these proteases, and both enzymes exhibited activity in the neutral to slightly basic pH range. Treatment of embryos with actinomycin D, an inhibitor of transcription, beginning up to 9 hr after fertilization, inhibited the subsequent appearances of the two proteases 48 hr after fertilization, as well as any morphological changes associated with gastrulation. Treatments beginning 15 and 21 hr after fertilization resulted in increased levels of proteases that correlate with arrests at successively more advanced stages of gastrulation. SDS‐PAGE zymographic analyses of five different embryo fractions indicated that the 57‐ and 50‐kDa proteases are localized in the blastocoel, and blastocoelic protease activity was further confirmed microscopically by in situ zymography. Hence, the 57‐ and 50‐kDa proteases are characterized as metalloproteases. Their expression is dependent on transcription of the embryonic genome, and their spatiotemporal appearance suggests an involvement in blastocoelic matrix remodeling during gastrulation. © 1995 Wiley‐Liss, Inc.