Biochimica et Biophysica Acta 1860 (2016) 1450–1465 Contents lists available at ScienceDirect Biochimica et Biophysica Acta journal homepage: www.elsevier.com/locate/bbagen Review Post-Translational Modifications in sperm Proteome: The Chemistry of Proteome diversifications in the Pathophysiology of male factor infertility Luna Samanta a,b, Nirlipta Swain b,AhmetAyaza, Vijay Venugopal a, Ashok Agarwal a,⁎ a American Center for Reproductive Medicine, Department of Urology, Cleveland Clinic, Cleveland, OH 44195, USA b Redox Biology Laboratory, Department of Zsoology, School of Life Sciences, Ravenshaw University, Cuttack – 753003, Odisha, India article info abstract Article history: Background: The spermatozoa undergo a series of changes in the epididymis to mature after their release from Received 22 October 2015 the testis and subsequently in the female reproductive tract after ejaculation to get capacitated and achieve fer- Received in revised form 26 March 2016 tilization potential. Despite having a silenced protein synthesis machinery, the dynamic change in protein profile Accepted 4 April 2016 of the spermatozoa is attributed either to acquisition of new proteins via vescicular transport or to several post- Available online 6 April 2016 translational modifications (PTMs) occurring on the already expressed protein complement. Scope of review: In this review emphasis is given on the PTMs already reported on the human sperm proteins Keywords: under normal and pathologic conditions with particular reference to sperm function such as motility and fertil- Spermatozoa Proteome ization. An attempt has been made to summarize different protocols and methods used for analysis of PTMs on Post-translational modifications sperm proteins and the newer trends those were emerging. Infertility Major conclusions: Deciphering the differential occurrence of PTM on protein at ultrastructural level would give Male factor us a better insight of structure-function relationship of the particular protein. Protein with multiple PTMs could be used to generate the complex interaction network involved in a physiological function of a sperm. It can be speculated that crosstalk between different PTMs occurring either on same/ other proteins actually regu- late the protein stability and activity both in physiological and pathological states. General significance: The analytical prospective of various PTMs reported in human spermatozoa and their rele- vance to sperm function particularly in various pathophysiological states, would pave way for development of biomarkers for diagnosis, prognosis and therapeutic intervention of male infertility. © 2016 Elsevier B.V. All rights reserved. 1. Introduction of defective post-translationally modified proteins and their substrates can lead to various human diseases and ailments. (See Table 1.) A proteome is the entire set of proteins that is or can be expressed by Mass spectrometry is one of the fundamental tools used to identify a cell, tissue, or organism at a given time. A cell proteome is much larger and analyze PTMs and quantify their changes. Modern proteomic and more complex than its respective encoding genome. Various factors approaches have made large-scale screening of complex mixtures of determine the structure and function of a cell proteome, including post- proteins possible, allowing us to precisely detect any chemical modifica- translational covalent modification of proteins. From a genetic point of tions. There are different branches of modification-specificproteomics view, any differences between the primary polypeptide sequence gen- based on the pattern of protein alteration–phosphoproteomics erated from translation and the final structure of a fully functional pro- for phosphorylation studies, diglycine remnant proteomics for tein are considered post-translational modification (PTM). These PTMs ubiquitination, degradomics for protein fragmentation, and redox pro- are chemical alterations in the protein structure typically catalyzed by teomics for oxidative damage to proteins [1–3]. exceedingly substrate-specific enzymes that regulate protein stability, Sperm essentially do not synthesize proteins. Thus, the dynamic na- distribution and function. Any failure to control such complex molecular ture of sperm proteome depends either on acquisition of new proteins processes could be detrimental or fatal to a cell because large amounts by vescicular transport or PTMs of already manufactured proteins. Therefore, PTMs on existing proteins play a pivotal role in regulating sperm function including maturation and acquisition of fertilizing potential. Of the 300 covalent modifications reported, phosphorylation, ⁎ Corresponding author at: Lerner College of Medicine, Andrology Center, American nitrosylation, glycosylation, methylation, lipid modification, Center for Reproductive Medicine, Cleveland Clinic, Mail Code X-11, 10681 Carnegie Avenue, Cleveland, OH, 44195, United States. ubiquitination, and acetylation are the most common [4]. Even under E-mail address: [email protected] (A. Agarwal). oxidative stress conditions, reactive oxygen species (ROS)-induced http://dx.doi.org/10.1016/j.bbagen.2016.04.001 0304-4165/© 2016 Elsevier B.V. All rights reserved. L. Samanta et al. / Biochimica et Biophysica Acta 1860 (2016) 1450–1465 1451 Table 1 A list of post-translational modifications of human sperm proteins and their role in sperm function. Sl. No. PTM Modified Protein in Human sperm Related function References 1 Phosphorylation Histone Sperm chromatin condensation [9,10] P1, P2 [13,19] Histone variants (TH2B, H2X, H1t) [20,21] [10,22], Bad, GSK-3β, HSP27, JNK/SAPK, mTOR, p38MAPK, p53 Sperm motility [30] AKAPs, F895 Capacitation [28,35,36] VCP, NSF, HSPA2 Zona binding [38] IZUMO1 Sperm-egg fusion [49] 2 Acetylation Histone Sperm chromatin condensation and epigenetic code [54,57,58,59] P1,P2 [19] 11 Tubulin proteins, ODF2, and Tektin proteins (TEKT3 and TEKT4) Sperm motility [58] 9 proteins of NADH dehydrogenase complex and 10 proteins of ATP synthase complex AKAP, CABYR, proteasome Capacitation [52,58] ACRBP, ACRV1, VDAC3 Acrosomal reaction [58] PDIA3/ERp57, ACE, ZPBP [52] RPL10A and RPL3 Ribosomal cytoplasmic protein [52] 3 Glycosylation SAGA-1 Sperm egg adhesion [74] GLIPR1L1, Acrosin, ADAM2 [64] IZUMO [75] GPX4 Antioxidant protection [64] 4 Nitrosylation HSPs, Tetkin, Tubulin, Semenoglein Sperm motility [82] Rys-R calcium channel Mobilization of Ca+2 stores in sperm [83,84] 5 Methylation Histone (H3) Epigenetic code [59] 6 Ubiquitination Histone (H2A, H2B) Histone replacement [179]. Prohibitin Marking defective spermatozoa for degradation [121] 7 SUMOylation Histone (H4) Sex chromosome silencing [127] SCP1, SCP2 Synaptonemal complex scaffold maintenance [122] RanGap1 Silencing gene translation [130,183] DRP1 Mitochondrial functioning [130] Topoisomerase IIα Chromatin condensation [183] Tubulins, ODF3 or SHIPPO, AKAP3 and AKAP4, Sperm motility [123] HSP70, HSP60, HSPgp96, GST, TCP1 complex proteins Stress related protein [123] RNE, SPAN-X, ubiquitin, VCP, Sperm maturation [123] Semenoglein Capacitation [123] protein modifications such as glutathionylation, nitro- tyrosine and special reference to their role in capacitation, motility, hyperactivation, dityrosine formation have been reported [5]. acrosome reaction, sperm-zona binding and egg activation [9]. The primary focus of this review is to provide an overview of the var- During epididymal transit, histone phosphorylation is a primary PTM ious PTMs reported in human spermatozoa and to discuss their rele- that regulates the chromatin remodeling process. Phosphorylation- vance to sperm function, particularly in various pathophysiological dephosphorylation of the sperm histone occurs during mammalian sper- states. In addition, we will also discuss the potential of using proteomic miogenesis and is most likely involved in the proper condensation of analysis to develop biomarkers for the diagnosis, prognosis and thera- spermatid chromatin [10,11]. Phosphorylation of specific histone variants peutic management of male infertility. may protect the chromatin from proteolytic degradation during the nucleohistone-to-nucleoprotamine transition and may later help in rapid activation of specific genes post fertilization [12,13].Furthermore, 2. Post-translational modifications (PTMs) in sperm phosphorylated protamine 1 (P1) and protamine 2 (P2) comprise a sig- nificant portion of the human sperm nuclei although they are present in 2.1. Phosphorylation Phosphorylation alters the structural conformation of a protein by introducing a covalently bound phosphate group to an amino acid (aa), especially tyrosine (tyr), serine (ser), and threonine (thr) of the polypeptide chain (Fig. 1). Because a charged and hydrophobic element gets added at the R group of the modified aa, nearby aa are affected as well, and it is these interactions that switch on or off the activity of a protein. In fact, the process of protein phosphorylation is regulated by the collaborative actions of protein kinases (PK) and protein phospha- tases (PP). The former catalyzes the addition of the phosphate group while the latter facilitates its removal. Phosphorylation is one of the major PTM controlling sperm function, especially epididymal maturation, motility, capacitation and acrosomal reaction. In fact, levels of tyr phosphorylation increase in specificcom- partments of human spermatozoon following capacitation and zona binding [6]. The sperm head is predominantly phosphorylated during early capacitation [7] followed by the