【摘要】：現(xiàn)代社會(huì)的發(fā)展為人們的生活提供了豐富的物質(zhì)保證，但同時(shí)人類的生殖健康卻因環(huán)境改變、生活節(jié)奏加快和壓力增大而面臨著巨大的考驗(yàn)。我國(guó)不孕不育率已經(jīng)占到育齡人群的10%，由于男性因素造成的不孕不育呈現(xiàn)明顯的上升趨勢(shì)。少精、弱精以及無精子癥是男性不育的主要原因，這個(gè)特殊的健康缺陷，成為導(dǎo)致家庭破裂、影響社會(huì)和諧的主要因素之一。因此，有關(guān)精子發(fā)生機(jī)理成為當(dāng)今生殖醫(yī)學(xué)領(lǐng)域中研究的熱點(diǎn)。 精子發(fā)生是一個(gè)高度復(fù)雜而有序的過程，從精原細(xì)胞增殖分化到精子成熟，需要經(jīng)歷有絲分裂、減數(shù)分裂和精子形成三個(gè)連續(xù)的階段。在這一過程中有許多基因?qū)ζ溥M(jìn)行調(diào)控，基因突變或表達(dá)異常均可導(dǎo)致不育。DAZ基因家族是導(dǎo)致男性不育的關(guān)鍵候選基因之一，也是近年來被廣泛研究的一個(gè)熱門基因家族。Boule基因是新發(fā)現(xiàn)的該基因家族成員，它可能調(diào)控減數(shù)分裂的發(fā)生，Boule基因的缺失、突變或者BOULE蛋白表達(dá)異常都可能導(dǎo)致精子發(fā)生過程阻滯，造成精子生成障礙，導(dǎo)致無精子癥而不育。 目前，，對(duì)Boule基因的生物學(xué)特征和作用機(jī)制缺乏深入全面的研究，本文利用生物信息學(xué)方法對(duì)人、倭黑猩猩、松鼠猴、牛、小鼠、虹鱒魚、斑胸草雀、黑腹果蠅、曼氏裂體吸蟲、擬南芥十個(gè)物種中Boule基因的核苷酸序列和蛋白序列進(jìn)行比較分析，利用不同物種蛋白序列構(gòu)建進(jìn)化樹，對(duì)其同源性和進(jìn)化關(guān)系進(jìn)行分析。利用生物信息學(xué)ExPASy軟件對(duì)其理化性質(zhì)、疏水性/親水性、信號(hào)肽、跨膜結(jié)構(gòu)、卷曲螺旋、細(xì)胞定位進(jìn)行分析，通過基于GOR算法的在線分析工具GOR4與PredictProtein對(duì)BOULE蛋白二級(jí)結(jié)構(gòu)、功能結(jié)構(gòu)域進(jìn)行預(yù)測(cè)和分析，根據(jù)Pfam數(shù)據(jù)庫(kù)對(duì)Boule功能以及信號(hào)通路進(jìn)行預(yù)測(cè)和分析。十個(gè)物種BOULE蛋白序列比較結(jié)果顯示，BOULE蛋白序列在物種進(jìn)化過程中變化較大，但是在哺乳動(dòng)物種間序列相似度達(dá)到90%~99%之間，具有較高的同源性。BOULE蛋白進(jìn)化樹分析表明，BOULE蛋白被聚為兩大類，擬南芥獨(dú)自為一類；其它物種為一類。BOULE蛋白生物學(xué)特征分析顯示：BOULE蛋白含有283個(gè)氨基酸，分子量為31.299KD，等電點(diǎn)為6.59，為酸性蛋白，屬于非分泌型蛋白，屬于非跨膜蛋白質(zhì)；BOULE蛋白存在于精原細(xì)胞的細(xì)胞質(zhì)，細(xì)胞核，線粒體中；二級(jí)結(jié)構(gòu)主要有α-螺旋(9.19%)、延伸鏈（22.26%）、無規(guī)則卷曲（68.55%）；結(jié)構(gòu)域分析結(jié)果表明BOULE蛋白含有RRM結(jié)構(gòu)域。對(duì)其功能預(yù)測(cè)分析顯示：BOULE蛋白與Pum2蛋白相互作用，調(diào)控細(xì)胞周期蛋白的轉(zhuǎn)錄；同時(shí)與減數(shù)分裂相關(guān)蛋白DAZ1、CDC25A、CDC25C相互作用，調(diào)控精原細(xì)胞的分化和減數(shù)分裂的發(fā)生，因此，Boule基因是精子細(xì)胞分化所必不可少的重要調(diào)控基因；本文通過生物信息學(xué)方法對(duì)BOULE蛋白進(jìn)化過程中的序列同源性、理化性質(zhì)及其在精子發(fā)生過程中的作用機(jī)理進(jìn)行分析和預(yù)測(cè)，有利于我們系統(tǒng)了解Boule基因在精子發(fā)生中的作用，為診斷和治療少精、弱精以及無精癥提供理論依據(jù)，在人類精子發(fā)生的研究中具有非常重要的意義。
[Abstract]:The development of modern society has provided abundant material guarantee for people's life, but at the same time, human reproductive health is facing a huge test because of environmental changes, quickened pace of life and increased pressure. Oligospermia, asthenospermia and azoospermia are the main causes of male infertility. This special health defect has become one of the main factors leading to family breakdown and affecting social harmony.
Spermatogenesis is a highly complex and orderly process, from spermatogonial proliferation and differentiation to sperm maturation, which involves three consecutive stages: mitosis, meiosis and spermatogenesis. Boule gene is a newly discovered member of this gene family. It may regulate meiosis. Boule gene deletion, mutation or abnormal expression of BOULE protein may lead to spermatogenesis arrest and spermatogenesis disorder. Lead to azoospermia and infertility.
At present, there is no thorough and comprehensive study on the biological characteristics and mechanism of Boule gene. The nucleotide and protein sequences of Boule gene in ten species of human, bonobo chimpanzee, squirrel monkey, cattle, mouse, rainbow trout, zebra finch, Drosophila melanogaster, Schistosoma mansoni and Arabidopsis thaliana were comparatively analyzed by bioinformatics. The phylogenetic tree was constructed by using different protein sequences and their homology and evolutionary relationships were analyzed. The physiochemical properties, hydrophobicity/hydrophilicity, signal peptide, transmembrane structure, coiled helix and cell localization of BOULE protein were analyzed by using bioinformatics software ExPASy. Boule function and signaling pathway were predicted and analyzed according to Pfam database. Comparison of BOULE protein sequences of ten species showed that BOULE protein sequences varied greatly during the evolution of species, but the similarity between mammalian species was between 90% and 99%. BOULE protein phylogenetic tree analysis showed that BOULE proteins were grouped into two major groups, Arabidopsis alone was one; other species were one. Biological characteristics of BOULE protein showed that BOULE protein contained 283 amino acids, molecular weight was 31.299 KD, isoelectric point was 6.59, it was acidic protein, belonged to non-secretory protein, belonged to non-transmembrane protein. BOULE protein was found in the cytoplasm, nucleus and mitochondria of spermatogonia; the secondary structure was mainly alpha-helix (9.19%), elongated chain (22.26%) and irregular convolution (68.55%); domain analysis showed that BOULE protein contained RRM domain. Cyclin transcription; interacts with meiosis-related proteins DAZ1, CDC25A and CDC25C to regulate the differentiation and meiosis of spermatogonia. Therefore, Boule gene is an essential regulatory gene for sperm cell differentiation. The analysis and prediction of physical and chemical properties and their mechanism in spermatogenesis will help us to understand the role of Boule gene in spermatogenesis systematically and provide theoretical basis for the diagnosis and treatment of oligospermia, asthenospermia and azoospermia.
【學(xué)位授予單位】：內(nèi)蒙古師范大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2014
【分類號(hào)】：Q811.4;R698.2

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