Hooper John D, Campagnolo Luisa, Goodarzi Goodarz, Truong Tony N, Stuhlmann Heidi, Quigley James P
Division of Vascular Biology, Department of Cell Biology, The Scripps Research Institute, 10550 North Torrey Pines Road, La Jolla, CA 92037, USA.
Biochem J. 2003 Aug 1;373(Pt 3):689-702. doi: 10.1042/BJ20030390.
We report the identification and characterization of mouse matriptase-2 (m-matriptase-2), an 811-amino-acid protein composed of an N-terminal cytoplasmic domain, a membrane-spanning domain, two CUB (complement protein subcomponents C1r/C1s, urchin embryonic growth factor and bone morphogenetic protein 1) domains, three LDLR (low-density-lipoprotein receptor class A) domains and a C-terminal serine-protease domain. All m-matriptase-2 protein domain boundaries corresponded with intron/exon junctions of the encoding gene, which spans approx. 29 kb and comprises 18 exons. Matriptase-2 is highly conserved in human, mouse and rat, with the rat matriptase-2 gene ( r-maltriptase-2 ) predicted to encode transmembrane and soluble isoforms. Western-blot analysis indicated that m-matriptase-2 migrates close to its theoretical molecular mass of 91 kDa, and immunofluorescence analysis was consistent with the proposed surface membrane localization of this protein. Reverse-transcription PCR and in-situ -hybridization analysis indicated that m-matriptase-2 expression overlaps with the distribution of mouse hepsin (m-hepsin, a cell-surface serine protease identified in hepatoma cells) in adult tissues and during embryonic development. In adult tissues both are expressed at highest levels in liver, kidney and uterus. During embryogenesis m-matriptase-2 expression peaked between days 12.5 and 15.5. m-hepsin expression was biphasic, with peaks at day 7.5 to 8.5 and again between days 12.5 and 15.5. In situ hybridization of embryonic tissues indicated abundant expression of both m-matriptase-2 and m-hepsin in the developing liver and at lower levels in developing pharyngo-tympanic tubes. While m-hepsin was detected in the residual embryonic yolk sac and with lower intensity in lung, heart, gastrointestinal tract, developing kidney tubules and epithelium of the oral cavity, m-matriptase-2 was absent in these tissues, but strongly expressed within the nasal cavity by olfactory epithelial cells. Mechanistic insight into the potential role of this new transmembrane serine protease is provided by its novel expression profile in embryonic and adult mouse.
我们报告了小鼠matriptase-2（m-matriptase-2）的鉴定和特性，这是一种由811个氨基酸组成的蛋白质，包含一个N端胞质结构域、一个跨膜结构域、两个CUB（补体蛋白亚成分C1r/C1s、海胆胚胎生长因子和骨形态发生蛋白1）结构域、三个LDLR（低密度脂蛋白受体A类）结构域和一个C端丝氨酸蛋白酶结构域。所有m-matriptase-2蛋白结构域边界与编码基因的内含子/外显子连接处相对应，该基因跨度约29 kb，包含18个外显子。Matriptase-2在人、小鼠和大鼠中高度保守，大鼠matriptase-2基因（r-maltriptase-2）预计编码跨膜和可溶性异构体。蛋白质免疫印迹分析表明，m-matriptase-2迁移接近其理论分子量91 kDa，免疫荧光分析与该蛋白在细胞膜表面定位的推测一致。逆转录PCR和原位杂交分析表明，m-matriptase-2的表达与成年组织及胚胎发育过程中小鼠hepsin（m-hepsin，一种在肝癌细胞中鉴定出的细胞表面丝氨酸蛋白酶）的分布重叠。在成年组织中，二者在肝脏、肾脏和子宫中的表达水平最高。在胚胎发育过程中，m-matriptase-2的表达在第12.5天至15.5天达到峰值。m-hepsin的表达呈双相性，在第7.5天至8.5天以及第12.5天至15.5天出现峰值。胚胎组织的原位杂交表明，m-matriptase-2和m-hepsin在发育中的肝脏中大量表达，在发育中的咽鼓管中表达水平较低。虽然在残留的胚胎卵黄囊中检测到m-hepsin，在肺、心脏、胃肠道、发育中的肾小管和口腔上皮中表达强度较低，但m-matriptase-2在这些组织中不存在，而是在鼻腔内由嗅觉上皮细胞强烈表达。该新跨膜丝氨酸蛋白酶在胚胎和成年小鼠中的新表达谱为深入了解其潜在作用机制提供了线索。
