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1.
Nematode sperm undergo a drastic physiological change during spermiogenesis (sperm activation). Unlike mammalian flagellated sperm, nematode sperm are amoeboid cells and their motility is driven by the dynamics of a cytoskeleton composed of major sperm protein (MSP) rather than actin found in other crawling cells. This review focuses on sperm from Caenorhabditis elegans and Ascaris suum to address the roles of external and internal factors that trigger sperm activation and power sperm motility. Nematode sperm can be activated in vitro by several factors, including Pronase and ionophores, and in vivo through the TRY-5 and SPE-8 pathways. Moreover, protease and protease inhibitors are crucial regulators of sperm maturation. MSP-based sperm motility involves a coupled process of protrusion and retraction, both of which have been reconstituted in vitro . Sperm motility is mediated by phosphorylation signals, as illustrated by identification of several key components (MPOP, MFPs and MPAK) in Ascaris and the characterization of GSP-3/4 in C. elegans . 相似文献
2.
Jiangli Dou Lianwan ChenYouqiao Hu Long Miao 《Biochimica et Biophysica Acta (BBA)/Molecular and Cell Biology of Lipids》2012,1821(6):934-942
Ejaculated mammalian sperm must acquire fertilization capacity after residing into the female reproductive tract, a process collectively known as capacitation. Cholesterol efflux was required for sperm maturation. Different from flagellated sperm, C. elegans sperm are crawling cells. C. elegans sperm are highly enriched with cholesterol though this animal species lacks biosynthetic pathway for cholesterol and its survival requires an exogenous cholesterol supply. The low abundance of cholesterol in C. elegans lipid extract is thought insufficient to form lipid microdomains ubiquitously in this organism. We present evidence that cholesterol is enriched in the plasma membrane of C. elegans spermatids and that cholesterol- and glycosphingolipids (GSLs)-enriched membrane microdomains (lipid microdomains) mediate sperm activation. Disruption of sperm lipid microdomains by acute manipulation of cholesterol in vitro blocks the sperm activation. Restriction of cholesterol uptake also results in the abnormal sperm activation in both males and hermaphrodites. Manipulation of the integrity of lipid microdomains by targeting the biosynthesis of GSLs inhibits sperm activation and the inhibition can be rescued by the addition of exogenous GSLs. The cleavage of glycosylphosphatidylinositol (GPI)-anchored proteins, which are exclusively found in lipid microdomains, also affects sperm activation. We conclude that localized signaling mediated by lipid microdomains is critical for worm sperm activation. Lipid microdomains composed of cholesterol and GSLs have been observed in flagellated sperm of several animal species, thus cholesterol, before its efflux from the plasma membrane, might be needed to assemble into a platform for some more important upstream signal sorting during spermatogenesis than was previously thought. 相似文献
3.
spe-12 encodes a sperm cell surface protein that promotes spermiogenesis in Caenorhabditis elegans 总被引:1,自引:0,他引:1
During spermiogenesis, Caenorhabditis elegans spermatids activate and mature into crawling spermatozoa without synthesizing new proteins. Mutations in the spe-12 gene block spermatid activation, rendering normally self-fertile hermaphrodites sterile. Mutant males, however, are fertile. Surprisingly, when mutant hermaphrodites mate with a male, their self-spermatids activate and form functional spermatozoa, presumably due to contact with male seminal fluid. Here we show that, in addition to its essential role in normal activation of hermaphrodite-derived spermatids, SPE-12 also plays a supplementary but nonessential role in mating-induced activation. We have identified the spe-12 gene, which encodes a novel protein containing a single transmembrane domain. spe-12 mRNA is expressed in the sperm-producing germ line and the protein localizes to the spermatid cell surface. We propose that SPE-12 functions downstream of both hermaphrodite- and male-derived activation signals in a spermatid signaling pathway that initiates spermiogenesis. 相似文献
4.
Juan J. Fraire‐Zamora Richard A. Cardullo 《Molecular reproduction and development》2010,77(9):739-750
Nematode spermatozoa are highly specialized amoeboid cells that must acquire motility through the extension of a single pseudopod. Despite morphological and molecular differences with flagellated spermatozoa (including a non‐actin‐based cytoskeleton), nematode sperm must also respond to cues present in the female reproductive tract that render them motile, thereby allowing them to locate and fertilize the egg. The factors that trigger pseudopod extension in vivo are unknown, although current models suggest the activation through proteases acting on the sperm surface resulting in a myriad of biochemical, physiological, and morphological changes. Compelling evidence shows that pseudopod extension is under the regulation of physiological events also observed in other eukaryotic cells (including flagellated sperm) that involve membrane rearrangements in response to extracellular cues that initiate various signal transduction pathways. An integrative approach to the study of nonflagellated spermatozoa will shed light on the identification of unique and conserved processes during fertilization among different taxa. Mol. Reprod. Dev. 77: 739–750, 2010. © 2010 Wiley‐Liss, Inc. 相似文献
5.
Spermiogenesis initiation in Caenorhabditis elegans involves a casein kinase 1 encoded by the spe-6 gene 总被引:1,自引:0,他引:1
Immature spermatids from Caenorhabditis elegans are stimulated by an external activation signal to reorganize their membranes and cytoskeleton to form crawling spermatozoa. This rapid maturation, termed spermiogenesis, occurs without any new gene expression. To better understand this signal transduction pathway, we isolated suppressors of a mutation in the spe-27 gene, which is part of the pathway. The suppressors bypass the requirement for spe-27, as well as three other genes that act in this pathway, spe-8, spe-12, and spe-29. Eighteen of the suppressor mutations are new alleles of spe-6, a previously identified gene required for an early stage of spermatogenesis. The original spe-6 mutations are loss-of-function alleles that prevent major sperm protein (MSP) assembly in the fibrous bodies of spermatocytes and arrest development in meiosis. We have isolated the spe-6 gene and find that it encodes a predicted protein-serine/threonine kinase in the casein kinase 1 family. The suppressor mutations appear to be reduction-of-function alleles. We propose a model whereby SPE-6, in addition to its early role in spermatocyte development, inhibits spermiogenesis until the activation signal is received. The activation signal is transduced through SPE-8, SPE-12, SPE-27, and SPE-29 to relieve SPE-6 repression, thus triggering the formation of crawling spermatozoa. 相似文献
6.
Kristin E. Fenker Angela A. HansenConrad A. Chong Molly C. JudBrittany A. Duffy J. Paul NortonJody M. Hansen Gillian M. Stanfield 《Developmental biology》2014
Motility of sperm is crucial for their directed migration to the egg. The acquisition and modulation of motility are regulated to ensure that sperm move when and where needed, thereby promoting reproductive success. One specific example of this phenomenon occurs during differentiation of the ameboid sperm of Caenorhabditis elegans as they activate from a round spermatid to a mature, crawling spermatozoon. Sperm activation is regulated by redundant pathways to occur at a specific time and place for each sex. Here, we report the identification of the solute carrier 6 (SLC6) transporter protein SNF-10 as a key regulator of C. elegans sperm activation in response to male protease activation signals. We find that SNF-10 is present in sperm and is required for activation by the male but not by the hermaphrodite. Loss of both snf-10 and a hermaphrodite activation factor render sperm completely insensitive to activation. Using in vitro assays, we find that snf-10 mutant sperm show a specific deficit in response to protease treatment but not to other activators. Prior to activation, SNF-10 is present in the plasma membrane, where it represents a strong candidate to receive signals that lead to subcellular morphogenesis. After activation, it shows polarized localization to the cell body region that is dependent on membrane fusions mediated by the dysferlin FER-1. Our discovery of snf-10 offers insight into the mechanisms differentially employed by the two sexes to accomplish the common goal of producing functional sperm, as well as how the physiology of nematode sperm may be regulated to control motility as it is in mammals. 相似文献
7.
During the process of spermiogenesis (sperm activation) in Caenorhabditis elegans, the dramatic morphological events that ultimately transform round sessile spermatids into polar motile spermatozoa occur without the synthesis of any new gene products. Previous studies have identified four genes (spe-8, spe-12, spe-27 and spe-29) that specifically block spermiogenesis and lead to hermaphrodite-specific fertility defects. Here, we report the cloning and characterization of a new component of the sperm activation pathway, spe-19, that is required for fertility in hermaphrodites. spe-19 is predicted to encode a novel single-pass transmembrane protein. The spe-19 mutant phenotype, genetic interactions and the molecular nature of the gene product suggest SPE-19 to be a candidate for the receptor/co-receptor necessary for the transduction of the activation signal across the sperm plasma membrane. 相似文献
8.
Juan J. Fraire-Zamora Gina Broitman-Maduro Morris Maduro Richard A. Cardullo 《International Journal of Biochemistry and Molecular Biology》2011,2(3):263-273
Nematode spermatozoa are highly specialized cells that lack flagella and, instead, extend a pseudopod to initiate motility. Crawling spermatozoa display classic features of amoeboid motility (e.g. protrusion of a pseudopod that attaches to the substrate and the assembly and disassembly of cytoskeletal filaments involved in cell traction and locomotion), however, cytoskeletal dynamics in these cells are powered exclusively by Major Sperm Protein (MSP) rather than actin and no other molecular motors have been identified. Thus, MSP-based motility is regarded as a simple locomotion machinery suitable for the study of plasma membrane protrusion and cell motility in general. This recent focus on MSP dynamics has increased the necessity of a standardized methodology to obtain C. elegans sperm extract that can be used in biochemical assays and proteomic analysis for comparative studies. In the present work we have modified a method to reproducibly obtain relative high amounts of proteins from C. elegans sperm extract. We show that these extracts share some of the properties observed in sperm extracts from the parasitic nematode Ascaris including Major Sperm Protein (MSP) precipitation and MSP fiber elongation. Using this method coupled to immunoblot detection, Mass Spectrometry identification, in silico prediction of functional domains and biochemical assays, our results indicate the presence of phosphorylation sites in MSP of Caenorhabditis elegans spermatozoa. 相似文献
9.
Ras, a member of the small G-protein family, regulates multiple signaling pathways in somatic cells. The objectives of the present study included the characterization and localization of Ras and the identification of its downstream effectors in hamster spermatozoa. Immunoblot analysis with a pan-Ras monoclonal antibody localized Ras to the particulate fraction of sonicated testicular and caput and cauda epididymal spermatozoa. However, Ras was present in both the particulate and soluble fractions of spermatocytes and round spermatids, suggesting that its membrane recruitment is completed during spermiogenesis. Immunoblots of plasma membrane fractions demonstrated that hamster spermatozoa express both N-Ras and K-Ras. Indirect immunofluorescence with pan-Ras antibody localized Ras to the flagellum. Immunoblot analysis of sperm plasma membrane fractions demonstrated the presence of phosphatidylinositol 3-kinase (PI3-kinase) and protein kinase C zeta (PKCzeta), the downstream targets of Ras, and coimmunoprecipitation analysis demonstrated their interaction with Ras. Inhibitors of PI3-kinase (wortmannin and 2-(4- morpholinyl)-8-phenyl-4H-1-benzopyran-4-one) and PKCzeta (staurosporine) inhibited the hyperactivation of sperm motility during capacitation in a dose-dependent manner, indicating that both PI3-kinase and PKCzeta are associated with development of this motility pattern. The interaction of Ras with both PI3-kinase and PKCzeta suggests that Ras may regulate several signaling pathways in spermatozoa. 相似文献
10.
Using an affinity-purified antibody to the major sperm protein (MSP) in Caenorhabditis elegans sperm we have shown by immunofluorescence that the MSP is localized in the fibrous bodies of spermatocytes and early spermatids, in the cytoplasm of late spermatids, and in the pseudopods of spermatozoa. The MSP can also form crystalline inclusions in mutant and wild-type sperm. The function of this protein is still unknown, but its ability to form filaments and its localization in the pseudopod, together with the lack of actin in these sperm suggest that the MSP may be required for amoeboid motility. 相似文献
11.
Northern pike (Esox lucius L.) spermatozoa are uniflagellated cells differentiated into a head without acrosome, a midpiece and a flagellar tail region flanked by a fin structure. Total, flagellar, head and midpiece lengths of spermatozoa were measured and show mean values of 34.5, 32.0, 1.32, 1.17 μm, respectively, with anterior and posterior widths of the midpiece measuring 0.8 and 0.6 μm, respectively. The osmolality of seminal plasma ranged from 228 to 350 mOsmol kg−1 (average: 283.88 ± 33.05). After triggering of sperm motility in very low osmolality medium (distilled water), blebs appeared along the flagellum. At later periods in the motility phase, the tip of the flagellum became curled into a loop shape which resulted in a shortening of the flagellum and a restriction of wave development to the proximal part (close to head). Spermatozoa velocity and percentage of motile spermatozoa decreased rapidly as a function of time postactivation and depended on the osmolality of activation media (P < 0.05). In general, the greatest percentage of motile spermatozoa and highest spermatozoa velocity were observed between 125 and 235 mOsmol kg−1. Osmolality above 375 mOsmol kg−1 inhibited the motility of spermatozoa. After triggering of sperm motility in activation media, beating waves propagated along the full length of flagella, while waves appeared dampened during later periods in the motility phase, and were absent at the end of the motility phase. By increasing osmolality, the velocity of spermatozoa reached the highest value while wave length, amplitude, number of waves and curvatures also were at their highest values. This study showed that sperm morphology can be used for fish classification. Sperm morphology, in particular, the flagellar part showed several changes during activation in distilled water. Sperm motility of pike is inhibited due to high osmolality in the seminal plasma. Osmolality of activation medium affects the percentage of motile sperm and spermatozoa velocity due to changes in flagellar wave parameters. 相似文献
12.
13.
Sperm storage within the oviductal isthmus prior to ovulation typically involves binding to oviductal epithelial cells, which are thought to modulate sperm functions including internal calcium concentration, membrane fluidity, and motility. Around the time of ovulation the spermatozoa are gradually released so that they eventually encounter the oocytes within the oviductal ampulla. Previous studies have shown that the oviductal epithelial cells selectively sequester high quality spermatozoa, but the role of oviductal fluid as a selective modulator of sperm function has been investigated to a lesser extent. Here we address the hypothesis that oviductal fluid is also likely to modulate sperm function. Using samples of porcine oviductal fluid collected in the follicular phase of the estrus cycle, we show that short exposure (20 min to 50 μg/mL of oviductal fluid proteins) to either of two separate proteins fractions (> or < 100 kDa) promotes boar sperm viability and acrosomal integrity, decreases sperm plasma membrane fluidity (measured using merocyanine S540), and increases zona binding and polyspermy during in vitro fertilization. Exposure to the lower molecular fraction significantly inhibited, but did not abolish, the bicarbonate-induced stimulation of motility. The results show that subpopulations of spermatozoa respond differentially to oviductal fluid, and suggest that exposure to oviductal fluid in vivo could exert a further level of functional sperm selection. 相似文献
14.
Zilli L Beirão J Schiavone R Herraez MP Cabrita E Storelli C Vilella S 《Theriogenology》2011,76(4):737-744
Our previous studies demonstrated that osmolality is the key signal in sperm motility activation in Sparus aurata spermatozoa. In particular, we have proposed that the hyper-osmotic shock triggers water efflux from spermatozoa via aquaporins. This water efflux determines the cell volume reduction and, in turn, the rise in the intracellular concentration of ions. This increase could lead to the activation of adenylyl cyclase and of the cAMP-signaling pathway, causing the phosphorylation of sperm proteins and then the initiation of sperm motility. This study confirms the important role of sea bream AQPs (Aqp1a and Aqp10b) in the beginning of sperm motility. In fact, when these proteins are inhibited by HgCl2, the phosphorylation of some proteins (174 kDa protein of head; 147, 97 and 33 kDa proteins of flagella), following the hyper-osmotic shock, was inhibited (totally or partially). However, our results also suggest that more than one transduction pathways could be activated when sea bream spermatozoa were ejaculated in seawater, since numerous proteins showed an HgCl2(AQPs)-independent phosphorylation state after motility activation. The role played by each different signal transduction pathways need to be clarified. 相似文献
15.
Mutations that disrupt the morphogenesis and localization of a sperm-specific organelle in Caenorhabditis elegans 总被引:4,自引:0,他引:4
Nematode sperm contain unusual organelles, membranous organelles, which undergo dramatic morphological changes during spermatogenesis. Early in spermatogenesis, the membranous organelle functions to transport sperm specific components to the spermatids; later, during the formation of the crawling spermatozoa, it adds new components to the cell surface as it fuses with the plasma membrane. Genetic analysis of spermatogenesis in the nematode Caenorhabditis elegans has revealed mutations that specifically disrupt the proper cellular localization and morphogenesis of this organelle. In animals homozygous for the either the known deficiency hcDf1 or the probable deficiency h12, the membranes of the membranous organelles are aberrantly covered with ribosomes. A mutation in the spermatogenesis-defective spe-10 gene causes severe defects in the morphogenesis of a fibrous body-membranous organelle complex. In both cases, these mutations also disrupt the proper localization of both nuclei and membranous organelles in haploid spermatids and spermatozoa. 相似文献
16.
Motility and cryopreservation of testicular sperm of European common frog, Rana temporaria were investigated. Collected testicular spermatozoa were immotile in solutions of high osmolalities: 300 mmol/l sucrose and motility inhibiting saline solution-MIS. Full sperm motility could be activated in distilled water or in a solution of 50 mmol/l NaCl, = 90 mosmol/kg, with 75-90% motility and 14-16 μm s−1 swimming velocity. Spermatozoa activated in distilled water and kept at room temperature ceased the motility within a period of 1 h. But when they were kept at 4 °C, no significant decrease in sperm motility and velocity occurred over a period of 1 h. Incubation of testicular sperm diluted 1:2 with MIS containing 10% DMSO, 5% glycerol, 10% methanol, or 10% propandiol for a period of 40 min at 4 °C showed that propandiol was the most toxic cryoprotectant for spermatozoa of European common frog R. temporaria. However, methanol was not toxic to spermatozoa during the 40 min incubation period, it failed to protect spermatozoa during the freezing and thawing process. DMSO and glycerol were useful penetrating cryoprotectants that interacted with sperm diluents in cryodiluent efficacy. In combination with the sucrose diluent, DMSO was a better cryoprotectant than glycerol, while in combination with MIS, DMSO and glycerol were similarly useful. Sperm was frozen at two freezing levels above the surface of liquid nitrogen. Sperm frozen 5 cm above the surface of liquid nitrogen resulted in immotile and non-viable spermatozoa. However, sperm frozen at 10 cm above the surface of liquid nitrogen showed 40-45% viability and 30-35% motility, compared to the untreated freshly collected testicular sperm. Addition of hen egg yolk had no positive effect on the post-thaw sperm motility, viability and hatching rate when added to sucrose cryodiluents. However, addition of 5% egg yolk to the MIS containing 5% glycerol and 2.5% sucrose significantly improved the hatching rate than all other treatments. Therefore, we conclude that, MIS and 300 mmol/l sucrose are suitable diluents for immotile storage of testicular semen. For cryopreservation, dilution to a final concentration of 5-6 × 106/ml in MIS with 5% glycerol, 2.5% sucrose and 5% egg yolk, frozen in liquid nitrogen vapour at 10 cm above its surface, and thawed at 22 °C for 40 s is a useful cryopreservation protocol for R. temporaria sperm. Further research is needed to determine the motility parameters and cryopreservation of spermatic urine of R. temporaria. 相似文献
17.
Motility of Pinctada margaritifera (Linnaeus, 1758); var: cumingii (Jameson, 1901) (P. margaritifera) spermatozoa collected from gonads are not immediately activated at spawning in seawater (SW) but motility occurs when spermatozoa are transferred into alkaline seawater (pH ranging from 9.0 to 11.4). This motility-activating effect of alkaline pH is reversed when pH is shifted back to more acidic values. In both cases, activity of sperm (% motile cells) increases gradually after alkaline pH activation then lasts for several minutes. The characteristics of these fully motile spermatozoa are described in details at the level of flagella: the wave amplitude and wave-length range 5 to 6 μm and 15 μm respectively, while the flagellar beat frequency is approximately 49 Hz. The velocity of sperm displacement is from 220 to 230 μm/sec. The general swimming pattern is almost circular: the head trajectories describe portions of circles intercalated with small linear segments. Spermatozoa saved in natural seawater at 4°C retain potent motility for several days and can be subsequently activated by alkaline seawater. Respiration and ATP concentration were measured in 3 conditions: regular seawater (pH 7.8), artificial diluent (pH 8.2), and alkaline Tris-buffered seawater (pH 10.5). Results show that sperm respiration rates are higher whereas ATP levels are lower in the latter two media. 相似文献
18.
Singaravelu G Chatterjee I Rahimi S Druzhinina MK Kang L Xu XZ Singson A 《Developmental biology》2012,365(2):376-383
Despite undergoing normal development and acquiring normal morphology and motility, mutations in spe-38 or trp-3/spe-41 cause identical phenotypes in Caenorhabditis elegans-mutant sperm fail to fertilize oocytes despite direct contact. SPE-38 is a novel, four-pass transmembrane protein and TRP-3/SPE-41 is a Ca(2+)-permeable channel. Localization of both of these proteins is confined to the membranous organelles (MOs) in undifferentiated spermatids. In mature spermatozoa, SPE-38 is localized to the pseudopod and TRP-3/SPE-41 is localized to the whole plasma membrane. Here we show that the dynamic redistribution of TRP-3/SPE-41 from MOs to the plasma membrane is dependent on SPE-38. In spe-38 mutant spermatozoa, TRP-3/SPE-41 is trapped within the MOs and fails to reach the cell surface despite MO fusion with the plasma membrane. Split-ubiquitin yeast-two-hybrid analyses revealed that the cell surface localization of TRP-3/SPE-41 is likely regulated by SPE-38 through a direct protein-protein interaction mechanism. We have identified sequences that influence the physical interaction between SPE-38 and TRP-3/SPE-41, and show that these sequences in SPE-38 are required for fertility in transgenic animals. Despite the mislocalization of TRP-3/SPE-41 in spe-38 mutant spermatozoa, ionomycin or thapsigargin induced influx of Ca(2+) remains unperturbed. This work reveals a new paradigm for the regulated surface localization of a Ca(2+)-permeable channel. 相似文献
19.
细胞运动、细胞迁移与细胞骨架研究进展 总被引:1,自引:0,他引:1
细胞定向运动与细胞骨架的动态循环密切相关。运动细胞在其伪足前沿依靠细胞骨架的不断聚合推动细胞膜的前进,在基部靠近细胞体部位通过细胞骨架的不断解聚收缩拖拉细胞体向前运动,细胞骨架的聚合与解聚通过伪足与支撑表面的吸附与解吸附而偶连。肌动蛋白组成的微丝骨架是大多数运动细胞的主要成分。外界刺激引起微丝细胞骨架动态变化的信号通路已逐步明了。线虫精子细胞的运动行为与阿米巴变形运动相似,但是在线虫精子细胞中没有肌动蛋白,而是以精子主要蛋白为基础形成细胞骨架驱动精子细胞的运动。与肌动蛋白不同,精子主要蛋白没有分子极性、ATP结合位点和马达蛋白。通过比较研究以上两种运动体系将有助于在分子水平上进一步阐明细胞运动的机理。 相似文献
20.
Changes in sperm features during the movement phase are especially interesting to study in external fertilization species whose sperm duration movement is long because this implies a significant adaptation of moving cells to the external medium. This study describes the changes in tetraploid Pacific oyster sperm characteristics in relation to time post activation.Sperm individually collected on three tetraploid males were activated in seawater. Their features were analysed over a 24 h period and compared to a sperm pool collected on three diploid males as a reference. The percentage of motile spermatozoa, the intracellular ATP content, and the fine structure of spermatozoa were studied in relation to time post activation. Furthermore, the fertilisation capacity of sperm individually collected on five diploid males was assessed after 1 and 24 h post activation.A forward progressive movement was maintained for at least a 20 h duration. Compared to diploid males, the percentage of motile spermatozoa was lower in tetraploid males. The intracellular ATP concentration was higher in spermatozoa from tetraploid males than in spermatozoa from diploid males. A decrease in ATP content was observed in the first 6 h post activation and severe alterations were observed in sperm morphology after 24 h. Then, a lower fertilisation capacity of sperm from diploid males was observed at the end of the movement phase.The cessation of Pacific oyster sperm motility was unlikely caused by ATP consumption as ATP concentration was still high at the end of sperm movement but rather caused by drastic changes in sperm morphology. Compared to sperm collected on diploid males, the lower quality of sperm from tetraploid males was emphasized by a shorter movement duration and deeper morphological alterations at the end of the movement phase. 相似文献