Elucidation of the protein composition of mouse seminal vesicle fluid

The seminal vesicles are male accessory sex glands that contribute the major portion of the seminal plasma in which mammalian spermatozoa are bathed during ejaculation. In addition to conveying sperm through the ejaculatory duct, seminal vesicle secretions support sperm survival after ejaculation, and influence the female reproductive tract to promote receptivity to pregnancy. Analysis of seminal vesicle fluid (SVF) composition by proteomics has proven challenging, due to its highly biased protein signature with a small subset of dominant proteins and the difficulty of solubilizing this viscous fluid. As such, publicly available proteomic datasets identify only 85 SVF proteins in total. To address this limitation, we report a new preparative methodology involving sequential solubilization of mouse SVF in guanidine hydrochloride, acetone precipitation, and analysis by label-free mass spectrometry. Using this strategy, we identified 126 SVF proteins, including 83 previously undetected in SVF. Members of the seminal vesicle secretory protein family were the most abundant, accounting for 79% of all peptide spectrum matches. Functional analysis identified inflammation and formation of the vaginal plug as the two most prominent biological processes. Other notable processes included modulation of sperm function and regulation of the female reproductive tract immune environment. Together, these findings provide a robust methodological framework for future SVF studies and identify novel proteins with potential to influence both male and female reproductive physiology.     

植物膜蛋白质组学研究技术现状

植物膜蛋白质组学是当前植物科学研究的热点领域。本文概论了蛋白质组学在植物膜蛋白研究中的应用,包括双向电泳前膜蛋白样品的制备以及植物质膜、液泡膜和其他膜蛋白组分的蛋白质组学研究进展,并介绍了植物膜蛋白质组学相关的数据库,最后对其发展作了展望。     

Proteomic analysis of human plasma and blood cells in nutritional studies: development of biomarkers to aid disease prevention

Proteomics is emerging as a valuable tool in nutritional research. Proteome analysis from plasma and blood cells can identify thousands of proteins that can potentially provide valuable new biomarkers for health, reveal early indications for disease disposition, assist in dietary responders from nonresponders, and enable the discovery of mechanisms of beneficial food component effects. This review discusses the latest developments in plasma, platelet and peripheral blood mononuclear cell proteomics, specifically in the field of nutritional proteomics, including issues relating to study design, sample preparation and data interpretation.     

Proteomic analysis of pikeperch seminal plasma provides novel insight into the testicular development of domesticated fish stocks

Control of the reproduction of domesticated stocks is considered a prerequisite for aquaculture development of pikeperch. However, knowledge about the physiology of the captive pikeperch male reproductive system and the biology of semen is very limited, especially regarding protein characteristics. The aims of our study were to characterize pikeperch sperm quantity and quality parameters and to analyze changes in the proteome of the same males spawned for the first and second times. Moreover, attempts were made to generate the first proteomic library of seminal plasma proteins. Semen collected during the first spawning season was characterized by lower sperm concentration and volume than for the second season. Using mass spectrometry-based label-free quantitative proteomics, we identified 850 proteins in the seminal plasma of pikeperch from both spawning seasons, and 65 seminal proteins were found to be differentially abundant between the first and second spawning seasons. The majority of differentially abundant proteins were involved in stress and immune responses, developmental processes, cofactor metabolic processes, proteolysis, cellular oxidant detoxification and organization of the extracellular matrix (ECM). In addition, several proteins unique to pikeperch seminal plasma were identified, including antifreeze proteins, hibernation-specific plasma proteins, lectins and vitellogenin. In summary, our results indicate that males that spawned for the first time were characterized by incompletely mature gonads and the expression of proteins associated with the early phase of spermatogenesis and ECM organization. On the other hand, males that spawned for the second time exhibited advanced gonadal maturation and expression of proteins related to the late stage of spermatogenesis and sperm maturation, including regulation of reactive oxygen species generation, bicarbonate production, sperm elongation and separation. The identification of a large number of seminal plasma proteins provides a valuable resource for understanding the functions of seminal plasma and the molecular mechanisms involved in testicular development and maturation in domesticated fish, which is a prerequisite for better control of reproduction in captivity.     

蛋白质组学及其技术发展

蛋白质组学产生于20世纪90年代,发展至今已日趋成熟。蛋白质组学是以生物体的全部或部分蛋白为研究对象,研究它们在生命活动过程中的作用、功能。蛋白质组学较之前的基因组学对于生命现象的解释更直接、更准确,近年得到了快速发展,并受到世界各国学者的高度关注。我们简要综述了蛋白质组学及其技术,并简单概述了这项技术在生命科学领域的应用。     

Development and application of proteomics technologies in Saccharomyces cerevisiae

Proteomics research focuses on the identification and quantification of "all" proteins present in cells, organisms or tissue. Proteomics is technically complicated because it encompasses the characterization and functional analysis of all proteins that are expressed by a genome. Moreover, because the expression levels of proteins strongly depend on complex regulatory systems, the proteome is highly dynamic. This review focuses on the two major proteomics methodologies, one based on 2D gel electrophoresis and the other based on liquid chromatography coupled to mass spectrometry. The recent developments of these methodologies and their application to quantitative proteomics are described. The model system Saccharomyces cerevisiae is considered to be the optimal vehicle for proteomics and we review studies investigating yeast adaptation to changes in (nutritional) environment.     

Proteomics of human seminal plasma: Identification of biomarker candidates for fertility and infertility and the evolution of technology

Proteomics is a research area that has developed rapidly in the last decade. It studies the large‐scale characterization of the full protein components of a cell, a tissue, or a biological fluid. In the last decade, clinical proteomics has developed new technology and bioinformatics useful in identifying molecular markers of pathology; the next decade might be the era of proteomics. Seminal plasma (SP) represents a good sample for proteomic analysis in the evaluation of male fertility/infertility. SP is an acellular fluid conglomerate, comprised of contributions from the epididymis and accessory sexual glands. Human SP contains many proteins that are important to the successful fertilization of the oocyte by the spermatozoa. Proteomic studies have identified numerous seminal‐specific proteins, and recent reports have provided a further understanding of their function with respect to male fertility. Upon further validation, these proteins may be useful in the clinical distinction between fertility and infertility. This article reviews the proteomic methods, such as one dimensional polyacrylamide gel electrophoresis (1D–PAGE), two‐dimensional polyacrylamide gel electrophoresis (2D–PAGE), and mass spectrometry (MS), employed to detect human SP markers involved in fertility and infertility. As such, proteomic studies will help the development of new techniques to identify novel biomarkers for a better clinical diagnosis and treatment of male infertility. Mol. Reprod. Dev. 80: 350–357, 2013. © 2013 Wiley Periodicals, Inc.     

Occurrence and reproductive roles of hormones in seminal plasma

Only 2–5% of seminal fluid is composed of spermatozoa, while the rest is seminal plasma. The seminal plasma is a rich cocktail of organic and inorganic compounds including hormones, serving as a source of nutrients for sperm development and maturation, protecting them from infection and enabling them to overcome the immunological and chemical environment of the female reproductive tract. In this review, a survey of the hormones found in human seminal plasma, with particular emphasis on reproductive hormones is provided. Their participation in fertilization is discussed including their indispensable role in ovum fertilization. The origin of individual hormones found in seminal plasma is discussed, along with differences in the concentrations in seminal plasma and blood plasma. A part of review is devoted to methods of measurement, emphasising particular instances in which they differ from measurement in blood plasma. These methods include separation techniques, overcoming the matrix effect and current ways for end-point measurement, focusing on so called hyphenated techniques as a combination of chromatographic separation and mass spectrometry. Finally, the informative value of their determination as markers of male fertility disorders (impaired spermatogenesis, abnormal sperm parameters, varicocele) is discussed, along with instances where measuring their levels in seminal plasma is preferable to measurement of levels in blood plasma.     

Proteomics in reproductive biology: Beacon for unraveling the molecular complexities

Proteomics, an interface of rapidly evolving advances in physics and biology, is rapidly developing and expanding its potential applications to molecular and cellular biology. Application of proteomics tools has contributed towards identification of relevant protein biomarkers that can potentially change the strategies for early diagnosis and treatment of several diseases. The emergence of powerful mass spectrometry-based proteomics technique has added a new dimension to the field of medical research in liver, heart diseases and certain forms of cancer. Most proteomics tools are also being used to study physiological and pathological events related to reproductive biology. There have been attempts to generate the proteomes of testes, sperm, seminal fluid, epididymis, oocyte, and endometrium from reproductive disease patients. Here, we have reviewed proteomics based investigations in humans over the last decade, which focus on delineating the mechanism underlying various reproductive events such as spermatogenesis, oogenesis, endometriosis, polycystic ovary syndrome, embryo development. The challenge is to harness new technologies like 2-DE, DIGE, MALDI-MS, SELDI-MS, MUDPIT, LC–MS etc., to a greater extent to develop widely applicable clinical tools in understanding molecular aspects of reproduction both in health and disease.     

蛋白质组学在附睾研究中的应用

蛋白质组学是在后基因组时代出现的一个新的研究领域,它从整体水平对蛋白质的表达和功能模式进行分析.附睾是精子成熟和贮存的场所,因其基因表达呈现出高度的区域特异性,可能成为蛋白质组学,尤其是差异蛋白质组学研究的一个理想模型.本文主要对不同物种的附睾蛋白质组以及附睾不同区域的差异表达蛋白的研究进展予以综述,期望为附睾功能研究及开发新型男性避孕药带来新的思路和方法.     

血浆蛋白质组学的研究进展

血浆蛋白质组学是研究血浆蛋白质的功能和变化的一门科学。血浆中蕴藏着生命机体的所有信息,因此只有彻底了解血浆中存在哪些蛋白质,才能知道如何利用血浆来预测人体对疾病的易感性并监控疾病的进程,以期达到对疾病进行早诊断早治疗。由于血浆蛋白质组动态范围大,给研究带来了很大的困难。尤其是高丰度蛋白质的存在影响了低丰度蛋白质的检测率。而低丰度蛋白质都是有意义的具有临床诊断价值的蛋白质。因此去除高丰度蛋白质的干扰成了血浆蛋白质组学研究的关键。近年来,血浆蛋白质组学相关研究技术也得到了长足进展,为深入研究血浆蛋白质做出了重要贡献。血浆蛋白质组学作为一种无创性的研究方法,值得我们去探讨。本文就血浆蛋白质组学研究进展情况做一简要综述。     

Proteomics reveals novel Drosophila seminal fluid proteins transferred at mating

Across diverse taxa, seminal fluid proteins (Sfps) transferred at mating affect the reproductive success of both sexes. Such reproductive proteins often evolve under positive selection between species; because of this rapid divergence, Sfps are hypothesized to play a role in speciation by contributing to reproductive isolation between populations. In Drosophila, individual Sfps have been characterized and are known to alter male sperm competitive ability and female post-mating behavior, but a proteomic-scale view of the transferred Sfps has been missing. Here we describe a novel proteomic method that uses whole-organism isotopic labeling to detect transferred Sfps in mated female D. melanogaster. We identified 63 proteins, which were previously unknown to function in reproduction, and confirmed the transfer of dozens of predicted Sfps. Relative quantification of protein abundance revealed that several of these novel Sfps are abundant in seminal fluid. Positive selection and tandem gene duplication are the prevailing forces of Sfp evolution, and comparative proteomics with additional species revealed lineage-specific changes in seminal fluid content. We also report a proteomic-based gene discovery method that uncovered 19 previously unannotated genes in D. melanogaster. Our results demonstrate an experimental method to identify transferred proteins in any system that is amenable to isotopic labeling, and they underscore the power of combining proteomic and evolutionary analyses to shed light on the complex process of Drosophila reproduction.     

Verification of male infertility biomarkers in seminal plasma by multiplex selected reaction monitoring assay

Seminal plasma is a promising biological fluid to use for noninvasive clinical diagnostics of male reproductive system disorders. To verify a list of prospective male infertility biomarkers, we developed a multiplex selected reaction monitoring assay and measured the relative abundance of 31 proteins in 30 seminal plasma samples from normal, nonobstructive azoospermia and post-vasectomy individuals. Median levels of some proteins were decreased by more than 100-fold in nonobstructive azoospermia or post-vasectomy samples, in comparison with normal samples. To follow up the most promising candidates and measure their concentrations in seminal plasma, heavy isotope-labeled internal standards were synthesized and used to reanalyze 20 proteins in the same set of samples. Concentrations of candidate proteins in normal seminal plasma were found in the range 0.1-1000 μg/ml but were significantly decreased in nonobstructive azoospermia and post-vasectomy. These data allowed us to select, for the first time, biomarkers to discriminate between normal, nonobstructive azoospermia, and post-vasectomy (simulated obstructive azoospermia) seminal plasma samples. Some testis-specific proteins (LDHC, TEX101, and SPAG11B) performed with absolute or nearly absolute specificities and sensitivities. Cell-specific classification of protein expression indicated that Sertoli or germ cell dysfunction, but not Leydig cell dysfunction, was observed in nonobstructive azoospermia seminal plasma. The proposed panel of biomarkers, pending further validation, could lead to a clinical assay that can eliminate the need for testicular biopsy to diagnose the category of male infertility, thus providing significant benefits to patients as well as decreased costs associated with the differential diagnosis of azoospermia.     

Identification and function of proteolysis regulators in seminal fluid

Proteins in the seminal fluid of animals with internal fertilization effect numerous responses in mated females that impact both male and female fertility. Among these proteins is the highly represented class of proteolysis regulators (proteases and their inhibitors). Though proteolysis regulators have now been identified in the seminal fluid of all animals in which proteomic studies of the seminal fluid have been conducted (as well as several other species in which they have not), a unified understanding of the importance of proteolysis to male fertilization success and other reproductive processes has not yet been achieved. In this review, we provide an overview of the identification of proteolysis regulators in the seminal fluid of humans and Drosophila melanogaster, the two species with the most comprehensively known seminal fluid proteomes. We also highlight reports demonstrating the functional significance of specific proteolysis regulators in reproductive and post‐mating processes. Finally, we make broad suggestions for the direction of future research into the roles of both active seminal fluid proteolysis regulators and their inactive homologs, another significant class of seminal fluid proteins. We hope that this review aids researchers in pursuing a coordinated study of the functional significance of proteolysis regulators in semen. Mol. Reprod. Dev. © 2012 Wiley Periodicals, Inc.     

水稻蛋白质组学研究进展

蛋白质组是一个基因组所表达蛋白质的总称,研究内容包括蛋白质基本氨基酸序列的鉴定到相关性状、修饰、功能及不同类型蛋白分子相互作用等等。本文简要介绍了蛋白质组学的产生背景,以及主要研究技术包括双向电泳(2D-PAGE)质谱、蛋白质芯片、酵母双杂交,并重点介绍了近期水稻蛋白质组学应用研究进展。     

Advances in fungal proteomics

Proteomics, the global analysis of proteins, will contribute greatly to our understanding of gene function in the post-genomic era. This review summarizes recent developments in fungal proteomics and also generalizes protocols for sample preparation from plant pathogenic fungi. Challenges and future perspectives of proteomics are discussed as well.     

A proteomics strategy for the analysis of bacterial biodegradation pathways

Bacterial biodegradation (bioremediation) is the use of microorganisms to break down organic materials into simpler compounds; it plays a pivotal role in the clean-up of hazardous wastes in the environment. Following the completion of genome sequencing in bacteria capable of biodegradation, functional genomic studies have played a major role in obtaining information on bacterial biodegradation pathways. Novel proteomics technologies have recently been developed to make it possible to analyze global protein expression. Proteomics can also provide important information on the life cycle, regulation, and post-translational modification of proteins induced under specific conditions. Proteomics technologies have been applied to the comprehensive study of bacterial biodegradation. In this paper, we introduce the proteomics technologies applicable to bacterial biodegradation studies, review the results of the proteomics analysis of representative biodegrading bacteria, and discuss the potential use of proteomics technologies in future biodegradation studies.     

蛋白质组学进展

在蛋白质水平上定量、动态、整体性研究生物体的蛋白质组学 ,将在后基因组时代大大增进我们对基因功能的理解。简要介绍了蛋白质组学的概念、研究手段 ,及最新进展     

Sixth Annual Meeting on Proteomic Sample Preparation,part of the second annual Getting Optimized Targets Summit

Atherosclerosis is a disease with higher levels of mortality in developed countries. Comprehension of the molecular mechanisms can yield very useful information in clinics for prevention, diagnosis and recovery monitoring. Proteomics represents an ideal methodology for this purpose, as proteins constitute the effectors of the different biological processes running during pathogenesis. To date, studies in atherosclerosis have been mainly focused on the search for plasma biomarkers. However, tissue proteomics allows going deeper into tissue secretomes, arterial layers or particular cells of interest, which, in turn, constitutes a more direct approximation to in vivo operating mechanisms. The aim of this review is to report latest advances in tissue proteomics in atherosclerosis and related diseases (e.g., aortic stenosis and ischemic injury).     

蛋白质组学的进展

蛋白质组学是在细胞的整体蛋白质水平上进行研究、从蛋白质整体活动的角度来认识生命活动规律的一门新学科.简要介绍蛋白质组学的科学背景及其最新发展.