Molecular Genetic Study of the Cystatin Gene Family

半胱氨酸蛋白酶抑制剂基因家族的分子遗传学研究

• 批准号: 02670842
• 负责人: SAITOH Eiichi
• 金额: $ 1.34万
• 依托单位: The Nippon Dental University at Niigata
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for General Scientific Research (C)
• 财政年份: 1990
• 资助国家: 日本
• 起止时间: 1990 至 1991
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-02670842/
• 关键词: Saliva; Cystatin; Molecular Cloning; Genomic Clone; Bowman-Birk Inhibitor; Molecular Evolution; DNA Sequence; Cysteine-Proteinase Inhibitor; シスタチンS; シスタチンD; 偽遺伝子; ホモロジ-; システインプロテア-ゼインヒビタ-; セリンプロテア-ゼインヒビタ-; 塩基配列; 分子遺伝学; ボ-マン・バ-クインヒビタ-; ドメイ

Human saliva contains a series of cysteine-proteinase inhibitors which are Classified into family II of "Cystatin Superfamily". Up to date, three molecular species of salivary (S-type) cystatins (cystatins S, SA and SN) have been elucidated. The physiological roles of these inhibitors could be the protection of the cells from inappropriate proteolysis and the regulation of cysteine proteinases both of host and bacterial origin. They share about 55% sequence homology with cystatin C, which is abundant in synovial fluid, seminal plasma and cerebrospinal fluid. A variant of cystatin C(Leu^<68> ->Gln) is known to deposit as amyloid fibrils in patients with hereditary cystatin C amyloid angiopathy.The synthesis of these four inhibitors is controlled by a multigene family leaving 6-7 members which is localized on human chromosome 20 - the cystatin gene family. From this gene family, five genes (named as the CST1, CST2, CST2B, CST3 and CST4) and two pseudogenes CSTP1 and CSTP2 (CST5) were, is … More olated and characterized. The three genes (CST1, CST2 and CST4), respectively, code for cystatins SN, SA and S. The CST2B is an allele at the CST2 locus. The CST3 codes for cystatin C. The S-type cystatin genes (CST1, CST2 and CST4) which contain the ATA and CAT boxes in their 5' -flanking regions, are differentially regulated and the expression of the genes is more restricted than of the CST3 gene, for cystatin C. In contrast to the S-type cystatin genes, the cystatin C gene shares some properties with the promoter of house keeping genes : lacking of typical CAT box and the presence of the binding sites of transcription factor Sp 1.The human cystatin genes sequenced here are composed of three exons encoding 76 or 81 (exon 1), 38 (exon 2) and 27 (exon 3) amino acids. These three-exon genes are closely related to an ancestral three-exon which generated contemporary nine exons coding for the kininogen heavy chain. Further analysis of the homology levels of the cDNA sequences of various proteinase inhibitors revealed that the second and third exons of the family II cystatin genes are significantly homologous with each other, and that DNA sequences of the two exons and exons 2, 3, 5, 6, 8 and 9 in the kininogen (family III cystatin) genes are extremely homologous to the CDNA sequencer encoding inhibitory domains of Bowman-Birk type serine-proteinase inhibitors. The kinetic study with synthetic peptides confirmed that the well conserved sequences in the protein domains encoded by the second and third exons of the family II cystatin gene possess inhibitory activities against the proteinase. Therefore it is evident that the exon-intron organization of the cystatin gene coincides with the structural and/or functional domains of the protein. These findings allowed us to conclude that the cystatin genes of families II and III have evolved by gene duplications from a common ancestral unit-length DNA sequence. Finally, we propose that cysteineproteinase inhibitors belonging to cystatin superfamily and serineproteinase inhibitors of Bowman-Birk family should be classified as the same family. Less

人唾液中含有一系列半胱氨酸蛋白酶抑制剂，属于Cystatin超家族中的第二家族。到目前为止，唾液中的三种半胱氨酸半胱氨酸氨基转移酶(S式)已被阐明(S、SA和SN)。这些抑制剂的生理作用可能是保护细胞免受不适当的蛋白分解，并调节宿主和细菌来源的半胱氨酸蛋白酶。它们与胱抑素C有55%的同源性，后者在滑液、精浆和脑脊液中含量丰富。胱抑素C的一个变异体(Leu^&lt；68&gt；-&gt；Gln)在遗传性胱抑素C淀粉样血管病变患者中以淀粉样纤维的形式沉积，这四种抑制物的合成受一个多基因家族的控制，该家族有6-7个成员，位于人类20号染色体上--cystatin基因家族。在这个基因家族中，有5个基因(命名为Cst 1、Cst 2、Cst 2B、Cst 3和Cst 4)和2个假基因Cst 1和Cst 2(Cst 5)是…更多的陈词滥调和特点。这三个基因(CST1、CST2和CST4)分别编码半胱氨酸蛋白SN、SA和S。CST2B是CST2基因座上的一个等位基因。对于Cystatin C，Cst3基因编码cystatin C。S型Cystatin基因(Cst1、Cst2和Cst4)的5‘侧翼区含有atA和CAT盒，它们的表达受到差异调控，并且表达受到比Cst3基因更大的限制。与S型cystatin基因相比，cystatin C基因与看家基因的启动子有一些共同的性质：缺乏典型的CAT盒，存在转录因子SP1的结合位点。38(外显子2)和27(外显子3)氨基酸。这些三外显子基因与祖先的三外显子密切相关，该三外显子产生了编码激肽原重链的当代九个外显子。进一步的同源性分析表明，家族II基因的第二和第三外显子之间存在显著的同源性，而激肽原基因(家族III)的两个外显子和外显子2、3、5、6、8和9的DNA序列与编码Bowman-Birk型丝氨酸蛋白酶抑制域的CDNA测序器高度同源。人工合成多肽的动力学研究证实，在第二家族胱抑素基因第二外显子和第三外显子编码的蛋白结构域中，保守的序列对该酶具有抑制活性。因此，很明显，cystatin基因的外显子-内含子组织与蛋白质的结构和/或功能域相一致。这些发现使我们可以得出结论，家族II和家族III的胱抑素基因是由共同的祖先单位长度DNA序列的基因复制进化而来的。最后，我们建议将属于胱抑素超家族的半胱氨酸蛋白酶抑制剂和Bowman-Birk家族的丝氨酸蛋白酶抑制剂归类为同一家族。较少

项目成果

Eiichi Saitoh et al.: "The human cystatin gene family:cloning of three members and evolutionary relationship between cystatins and BowmanーBirk type proteinase inhibitors" Biomedica and Biochimica Acts. (1991)

Eiichi Saitoh 等人：“人类半胱氨酸蛋白酶抑制剂基因家族：三个成员的克隆以及半胱氨酸蛋白酶抑制剂和 Bowman-Birk 型蛋白酶抑制剂之间的进化关系”Biomedica 和 Biochimica Acts（1991）。

Isemura, S., Saitoh, E., Sanada, K. and Minakata, K.: "Identification of Full-Sized Forms of Salivary (S-Type) Cystatins (Cystatin SN, Cystatin SA, Cystatin S, and Two Phosphorylated Forms of Cystatin S) in Human Whole Saliva and Determination of Phosphor

Isemura, S.、Saitoh, E.、Sanada, K. 和 Minakata, K.：“全尺寸形式唾液（S 型）胱抑素（胱抑素 SN、胱抑素 SA、胱抑素 S 和两种磷酸化形式）的鉴定

Satoko Isemura,Eiichi Saitoh,Kazuo Sanada,Kayoko Minakata: "Identification of Fullーsized Forms of Salivary(SーType)Cystatins(Cystatin SN,Cystatin SA,Cystatin S,and Two Phosphorylated Forms of Cystatin S)in Human Saliva and Determination of Phosphorylation

Satoko Isemura、Eiichi Saitoh、Kazuo Sanada、Kayoko Minakata：“人类唾液中全尺寸唾液（S 型）胱抑素（胱抑素 SN、胱抑素 SA、胱抑素 S 和两种磷酸化形式的胱抑素 S）的鉴定和测定磷酸化

Eiichi Saitoh Satoko Isemura Kazuo Sanada Koji Ohnishi: "The Human Cystatin Gene Family: Cloning of Three Members and Evolutionary Relationship between Cystatins and Bowman-Birk Type Proteinase Inhibitors." Biomedica Biochimica Acta. 50. 599-605 (1991)

Eiichi Saitoh Satoko Isemura Kazuo Sanada Koji Ohnishi：“人类半胱氨酸蛋白酶抑制剂基因家族：三个成员的克隆以及半胱氨酸蛋白酶抑制剂和鲍曼-伯克型蛋白酶抑制剂之间的进化关系。”

Eiichi Saitoh et al.: "Cystatins of Family II are harboring two domains which retain inhibitory activities against the proteinases" Biochemical and Biophysical Research Communications. (1991)

Eiichi Saitoh 等人：“II 家族的半胱氨酸蛋白酶抑制剂具有两个保留对蛋白酶的抑制活性的结构域”《生物化学和生物物理研究通讯》。