近日,，北京生命科學(xué)研究所董夢(mèng)秋實(shí)驗(yàn)室,、中科院生物物理所苗龍實(shí)驗(yàn)室以及計(jì)算所pFind研究組在國(guó)際著名雜志PNAS上在線發(fā)表題為 “Nematode sperm maturation triggered by protease involves sperm-secreted serine protease inhibitor (Serpin)”的論文，文章揭示了線蟲精細(xì)胞的激活機(jī)制和精子競(jìng)爭(zhēng)的機(jī)制,。
線蟲綱（nematode）物種包括雌雄同體的秀麗線蟲（C. elegans）和雌雄異體的豬蛔蟲（Ascaris suum）的精細(xì)胞在形成之后需要被激活,，從圓形的,、不能游動(dòng)的精細(xì)胞變成用偽足爬行的、有受精能力的精子,。未活化的精細(xì)胞體內(nèi)有大量的叫做membranous organelles (MO) 的小泡,。小泡的內(nèi)容物在精子活化過程中通過胞吐（exocytosis）釋放到細(xì)胞外。如果胞吐被抑制,，精細(xì)胞則不能被活化,。究竟什么分子被MO包裹，然后釋放到胞外,，它們的功能是什么,，一直是未解之謎。在這篇文章中,，作者通過多種實(shí)驗(yàn)手段,，包括經(jīng)典的生物學(xué)純化和一種新的基于質(zhì)譜的肽段從頭測(cè)序方法，發(fā)現(xiàn)豬蛔蟲的精細(xì)胞在活化過程中釋放出一個(gè)絲氨酸蛋白酶的抑制因子（命名為As_SRP-1）,。進(jìn)一步研究發(fā)現(xiàn)該蛋白被排放后執(zhí)行了兩種功能：（1）As_SRP-1結(jié)合到已經(jīng)活化或開始活化的精子表面,，維持偽足細(xì)胞骨架的動(dòng)態(tài)組裝；如果As_SRP-1的結(jié)合被破壞,，精子的運(yùn)動(dòng)能力便隨之喪失,。（2）As_SRP-1抑制一個(gè)由輸精管末端分泌的、活化精細(xì)胞的絲氨酸蛋白酶,。因該蛋白酶與胰蛋白酶（Trypsin）相似,，被命名為As_TRY-5。通過抑制As_TRY-5,，As_SRP-1阻礙周圍的精細(xì)胞被活化,。這可能是豬蛔蟲精子競(jìng)爭(zhēng)的一種方式。現(xiàn)有的遺傳學(xué)證據(jù)提示,，通過蛋白酶和蛋白酶抑制因子調(diào)節(jié)精子活化或精子競(jìng)爭(zhēng)的機(jī)制在秀麗線蟲中很可能是保守的,。另外，其它動(dòng)物比如小鼠的精液里也有蛋白酶和蛋白酶抑制劑,，所以類似的調(diào)節(jié)機(jī)制可能廣泛存在,。
這篇文章的一個(gè)技術(shù)亮點(diǎn)是不依賴于蛋白數(shù)據(jù)庫(kù)的肽段從頭測(cè)序（de novo sequencing）。以前的從頭測(cè)序效果欠佳,，鮮有實(shí)際應(yīng)用,。為了此項(xiàng)研究的需要，董夢(mèng)秋實(shí)驗(yàn)室與中科院計(jì)算所賀思敏研究組共同開發(fā)了肽段從頭測(cè)序的質(zhì)譜實(shí)驗(yàn)流程和算法,，達(dá)到了與數(shù)據(jù)庫(kù)搜索相當(dāng)?shù)蔫b定效果（Chi H. et al.,， J. Proteome Res. 2010）。本文中的兩個(gè)蛋白As_SRP-1和As_TRY-5都是用肽段從頭測(cè)序技術(shù)pNovo鑒定到的,，表明pNovo足以支持生物學(xué)研究,，尤其是對(duì)于新蛋白（序列未被收納到現(xiàn)有數(shù)據(jù)庫(kù)中）的鑒定有極大的幫助,。
生物物理所趙艷梅博士是本文第一作者。董夢(mèng)秋博士與生物物理所苗龍博士為本文的共同通訊作者,。參與本研究工作的還有我所董夢(mèng)秋實(shí)驗(yàn)室的博士生張攀,、宋春青、和杜立林實(shí)驗(yàn)室的張美??；中科院計(jì)算所賀思敏教授和博士生遲浩；生物物理所的孫維,、馬軒,、尚云龍、汪斌,、胡攸喬,、孟凡霞；ThermoFisher Scientific的Dr. Zhiqi Hao和Dr. Andreas F. Huhmere,；以及Emory University的Dr. Steven W. L’Hernault,。杜立林博士指導(dǎo)了肽段的BLAST search?？萍疾?、國(guó)家自然科學(xué)基金委員會(huì)、中科院,、和北京市政府提供了科研資助,。（生物谷Bioon.com）
doi:10.1073/pnas.1109912109
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Nematode sperm maturation triggered by protease involves sperm-secreted serine protease inhibitor (Serpin)
Yanmei Zhaoa, Wei Suna,b,1, Pan Zhangc,1, Hao Chib,d,1, Mei-Jun Zhangc,1, Chun-Qing Songc, Xuan Maa, Yunlong Shanga,b, Bin Wanga, Youqiao Hua,b, Zhiqi Haoe, Andreas F. Hühmere, Fanxia Menga, Steven W. L'Hernaultf, Si-Min Hed, Meng-Qiu Dongc,2, and Long Miaoa,2
Spermiogenesis is a series of poorly understood morphological, physiological and biochemical processes that occur during the transition of immotile spermatids into motile, fertilization-competent spermatozoa. Here, we identified a Serpin (serine protease inhibitor) family protein (As_SRP-1) that is secreted from spermatids during nematode Ascaris suum spermiogenesis (also called sperm activation) and we showed that As_SRP-1 has two major functions. First, As_SRP-1 functions in cis to support major sperm protein (MSP)-based cytoskeletal assembly in the spermatid that releases it, thereby facilitating sperm motility acquisition. Second, As_SRP-1 released from an activated sperm inhibits, in trans, the activation of surrounding spermatids by inhibiting vas deferens-derived As_TRY-5, a trypsin-like serine protease necessary for sperm activation. Because vesicular exocytosis is necessary to create fertilization-competent sperm in many animal species, components released during this process might be more important modulators of the physiology and behavior of surrounding sperm than was previously appreciated.
