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The proper functioning and assembly of the sperm flagella structures contribute significantly to spermatozoa motility and overall male fertility. However, the fine mechanisms of assembly steps are poorly studied due to the high diversity of cell types, low solubility of the corresponding protein structures, and high tissue and cell specificity. One of the open questions for investigation is the attachment of longitudinal columns to the doublets 3 and 8 of axonemal microtubules through the outer dense fibers. A number of mutations affecting the assembly of flagella in model organisms are known. Additionally, evolutionary genomics data and comparative analysis of flagella morphology are available for a set of non-model species. This review is devoted to the analysis of diverse ultrastructures of sperm flagellum of Metazoa combined with an overview of the evolutionary distribution and function of the mammalian fibrous sheath proteins. [ABSTRACT FROM AUTHOR]

CABYR is a protein that regulates calcium-binding tyrosine phosphorylation located in sperm flagella, which increases significantly during sperm capacitation and is primarily involved in sperm capacitation and motility. This study aimed to analyze the expression regulation pattern, sequence characteristics and potential biological function of CABYR gene in Banna mini-pig inbred line (BMI). The testes of adult BMI boars were used for RNA-seq; the complete coding sequence of CABYR was cloned; the sequence, structural characteristics, interacting proteins, KEGG and GO of CABYR were analyzed; the ceRNA regulatory network of CABYR was built using RNA-seq data. The average expression level and TPM value of CABYR gene obtained by RNA-seq was 11 187.50 and 225.29, respectively. The full-length CDS of CABYR was 1149 bp (GenBank accession number: OK042309) encoding 382 amino acids (GenBank login number: UYO37316). The amino acid sequence alignment analysis of multiple species revealed that the similarity between BMI and other species exceeded 73%, indicating a relatively conserved sequence. The CABYR protein was found to contain the DD_CABYR_SP17 conserved domain, the results of species phylogenetic tree analysis met the clustering criteria. Other analyses, including protein-protein interaction (PPI) networks, KEGG, and GO, revealed that BMI CABYR interacts with 50 proteins involved in a variety of functions, such as sexual reproduction and reproductive processes. CABYR was involved in 7 GOs, including five cellular components, one molecular functions, and one biological processes, according to functional annotation. Six miRNAs were found to regulate the CABYR gene via targeted interactions. Additionally, 8, 6, and 1 lncRNAs were found to interact with ssc-miR-4331-3p, ssc-miR-744, and ssc-miR-491, respectively, in association with CABYR. This study examines the expression of CABYR in the testis of BMI, as well as its molecular structure characteristics and expression regulatory network. These findings provide a foundation for future research on the involvement of the CABYR gene in spermatogenesis processes in BMI pigs, particularly in relation to crucial biological processes such as sperm motility and fibrous sheath development. [ABSTRACT FROM AUTHOR]

Purpose: The number of accessible central veins (CVs) affects the prognosis of patients with intestinal failure (IF). The loss of residual CVs should be avoided. We, therefore, evaluated the efficacy of a new CV catheter-exchange technique using a subcutaneous fibrous sheath (FS) in pediatric IF patients. Methods: We retrospectively collected the CV catheter (CVC) data of pediatric IF patients managed from January 2009 to December 2019. The data were divided into two groups; Groups 1 (CVCs placed with the FS method) and Group 2 (CVCs placed by the primary or another insertion). The main outcome was the CVC indwelling time. Results: Eighty-five CVCs were analyzed. The FS method was attempted in 47 cases and succeeded in 40 (85%). No significant difference was observed between the groups regarding characteristics. A log-rank test revealed an equivalent CVC indwelling time between the two groups (Group 1: 268 [126–588] days vs. Group 2: 229 [126–387] days, p = 0.256). Conclusions: The FS method is highly recommended for pediatric IF patients, as its attempt showed a high success rate with an indwelling time equivalent to primary insertion. The FS method leads to the prolonged use of a single CV and thereby contributes to improving the outcomes of pediatric IF patients. [ABSTRACT FROM AUTHOR]

The calcium-binding protein spermatid-associated 1 (CABS1) localizes to the principal piece of mature sperm flagella. Deletion of CABS1 results in subfertility in male mice, possibly due to an impaired annulus in the sperm flagella. However, it is unknown whether there are other mechanisms by which CABS1 affects male fertility. Our current investigation has uncovered that CABS1 is located in the midsection of the flagellum in testicular sperm and the principal piece in epididymal sperm. Moreover, male mice lacking CABS1 exhibit a defect in the progressive motility of sperm. Furthermore, the regulation of calcium levels, which has been reported to have a significant impact on sperm motility, capacitation, and the acrosome reaction, is also affected when sperm are exposed to alkalized high-salt buffer (pH 8.0) and progesterone (100 μM) in Cabs1-null spermatozoa. This alteration in calcium response may contribute to changes in the phosphorylation of PKA substrates and subsequent phosphorylation of tyrosine residues. Additionally, the absence of CABS1 leads to a defective fibrous sheath and abnormal configuration of doublet microtubules in post-testicular sperm. These findings indicate that the absence of CABS1 also disrupts the structural integrity of the fibrous sheath, resulting in male subfertility. The highly conserved nature of CABS1 in humans suggests that it could potentially be a contributing factor to asthenozoospermia in men., (© 2024 Wiley Periodicals LLC.)

Immunity-related GTPases (IRGs), also known as p47 GTPases, are a family of interferon-inducible proteins that play roles in immunity defense against intracellular pathogens. Although the molecular functions of IRGs have been well studied, the function of the family member, IRGC1, remains unclear. IRGC1 is unique among IRGs because its expression is not induced by interferon and it is expressed predominantly in the testis. Further, IRGC1 is well conserved in mammals unlike other IRGs. Here, we knocked out (KO) Irgc1 in mice using the CRISPR/Cas9 system and found that the fertility of Irgc1 KO males was severely impaired because of abnormal sperm motility. Further analyses with a transmission electron microscope revealed that the fibrous sheath (FS), an accessory structure of the sperm tail, was disorganized in Irgc1 KO mice. In addition, IRGC1 was detected in the sperm tail and fractionated with FS proteins. These results suggest that IRGC1 is a component of the FS and is involved in the correct formation of the FS. [Display omitted] • IRGC1, a testis-enriched immunity related GTPase, is important for male fertility in mice. • Irgc1 KO mice exhibited impaired sperm motility. • Fibrous sheath structures were disrupted in Irgc1 KO spermatozoa. • IRGC1 fractionates with fibrous sheath proteins, suggesting that IRGC1 is a component of the fibrous sheath. [ABSTRACT FROM AUTHOR]

Objective To evaluate the value of CABYR protein and its complex with FSCB in maintaining sperm fertilization ability and investigate the possible mechanism of physiological function of the coruplex by observing the natural fertility of Fscb/Cabyr double gene knockout and Cabyr single gene knockout male mice. Methods Fscb/Cabyr (--/--) double gene knockout male mice, Fscb/Cabyr (++/--) single gene knockout male mice and wild background male mice were mated with wild-type female mice respectively, and the pregnancy and litter production of female mice in each group were observed. The sperm of male mice with different genotypes were labeled with Fluo-4 AM calcium fluorescent probe in vitro and observed under fluorescence microscope, and the level of calcium ion in the sperm were detected by ELISA; The expression of CATSPER channel proteins in sperm of mice with different genotypes was measured by Western blotting. Results All the male mice with Fscb/Cabyr (--/--) double gene knockout and Fscb/Cabyr (++/--) single gene knockout lost their reproductive capacity completely, and failed to make any female mice pregnant and give birth. Compared with the wild-type male mice, the sperm calcium ion concentration of Fscb/Cabyr (--/--) and Fscb/Cabyr (++/--) male mice was decreased significantly (P < 0.000 1); The expression of CATSPER-1 protein in the sperm of Fscb/Cabyr(--/--) mice was decreased obviously. The expression of CATSPER-4 in the sperm of Fscb/Cabyr(--/--) and Fscb/Cabyr(++/--) mice were notably decreased. Conclusion The deletion of either CABYR or FSCB/CABYR protein complex in male sperm leads to the loss of reproductive function in the male mice, which may be realized by affecting the expression of calcium channel proteins CATSPER-1 and CATSPER-4, concentration of calcium ions in sperm, and then motility function of sperm.

Objective To investigate the changes in morphological characteristics of sperm flagellum and in sperm motility parameters in Fscb/Cabyr double gene knockout (Fscb/Cabyr--/--) mice. Methods The testis and epididymis tissues of male mice with different genotypes were collected and the organ-to-body weight ratio was calculated. The changes of spermatogenesis function in the testicular tissues of the mice were observed using HE staining. The parameters of sperm motility were analyzed using a computer-aided sperm analysis (CASA) system, and sperm capacitation and acrosome reaction were observed with in vitro culture. The morphological characteristics of the sperms under optical microscope and electron microscope were compared among the mice with different genotypes. Results No significant difference was found in sperm density in the epididymis among the mice with 5 different genotypes. The progressive rate (PR) and motility rate (PR+NP), mean curvilinear velocity (VCL), mean straight-line velocity (VSL), mean path velocity (VAP), beat cross frequency (BCF) and amplitude of lateral head displacement (ALH) of the sperms were significantly lower in Fscb/Cabyr--/-- and Fscb/Cabyr++/-- mice than in the wild-type mice (P < 0.01). The Fscb/Cabyr--/-- and Fscb/Cabyr++/-- mice showed significantly lowered percentages of normal sperms 1.45)% and (53.22±3.51)%] as compared with the wild-type mice and significantly higher rates of sperm tail folding] than the wild-type mice. Ultrastructural observation of the sperm flagellum in Fscb/Cabyr--/-- and Fscb/Cabyr++/-- mice revealed that the longitudinal columns of the fibrous sheath disappeared in most of the sperms with disorganized circumferential ribs and heterogeneous reduction of the electron density in the sperms; some sperms showed disruption of the axoneme structure in the flagella with occasional disappearance and dissociation of the doublet microtubules at the center. Conclusion Both Cabyr single gene knockout and Fscb/Cabyr double gene knockout can cause significant abnormalities in flagellum morphology and motility of the sperms in male mice. CABYR protein is essential for maintaining normal fibrous sheath structure and normal motility of the sperm flagella.

Abstract Background Sperm morphology mainly refers to the shape of the head, the length of the flagellar segments, including the midpiece, principal piece and end piece, and the size of the accessory structures, including axonemes, outer dense fibers (ODFs), mitochondrial sheath (MS) and fibrous sheath (FS). Across species, there is considerable diversity in morphology. An established theory posits that the length of the sperm flagellum, especially the length of the midpiece, is a critical factor influencing sperm metabolism and velocity. However, our understanding of the relationships between sperm ultrastructures and the sperm flagellar length is incomplete. Methods The morphologies of sperm from 10 mammalian species, human, mouse, rat, dog, rabbit, goat, pig, bull, guinea pig and golden hamster, were examined by scanning electron microscopy (SEM) and transmission electron microscopy (TEM). According to the SEM and TME images, the length of sperm heads and flagellar segments, the cross-sectional areas of the accessory structures and flagella and the width of sperm heads were measured using Image J software. The variation tendencies (referred to as slope) of the accessory structures along flagella were calculated by the linear regression method. Mitochondrial functions were measured using commercial kits. The velocities of sperm were measured using CASA software. Results The three-dimensional morphologies of sperm from 10 species and the slopes of internal accessory structures along flagella were obtained. The width of the axoneme tapered slightly from the base to the tip of the sperm flagellum, and slopes of the axonemes correlated negatively with the variability in flagellar length across species. Additionally, the cross-sectional areas of the ODFs and/or the MS were positively correlated with the lengths of the midpiece, principal piece, and total flagellum, as well as with sperm velocities. Mitochondrial volumes were positively correlated with ATP content and sperm swimming velocities. Conclusions Our results not only show the relationship between sperm internal structures, flagellar length and sperm physiology but also provide sizes of mitochondria and ODFs as new targets with which to study the regulation of sperm length and velocity.

Résumé Contexte Les limites des analyses conventionnelles du sperme ont mis en évidence la nécessité de moyens supplémentaires d’évaluation de la qualité du sperme. Matériel et Méthodes La méthode d’électrophorèse unidimensionnelle a été utilisée pour évaluer la teneur et la qualité protéique dans des échantillons d’éjaculats individuels de 245 hommes avec des paramètres spermatiques connus. Résultats La teneur en protéines du sperme variait d’un échantillon à l’autre, en particulier dans la gamme des poids moléculaires élevés. L’intensité des bandes de 80–110 kDa était corrélée à la mobilité progressive (r = 0,15, p = 0,015) et était significativement plus élevée (p = 0,0367) dans le groupe des hommes avec des paramètres conventionnels spermatiques supérieurs aux valeurs de référence (OMS, 2010) comparé au groupe d’hommes dont l’éjaculat présentait au moins un paramètre spermatique (volume de sperme, concentration de spermatozoïdes, numération des spermatozoïdes, motilité progressive, formes normales et indice d’anomalies multiples) en deça de la valeur de référence. Les bandes dans la gamme de 80–110 kDa ont été identifiées en spectrométrie de masse et se sont révélées être des protéines de la gaine fibreuse du flagelle: la protéine d’ancrage A-kinase 4, la protéine d’ancrage A-kinase 3 et l’hexokinase de type 1. Conclusion La méthode d’électrophorèse unidimensionnelle constitue une méthode simple, rapide, fiable et peu coûteuse pour analyser la qualité des protéines du sperme dans les éjaculats humains individuels.

The development and maintenance of the correct morphology of sperm is important for their functions. Cellular morphogenesis of sperm occurs during the post-meiotic developmental stage; however, little is known about what coordinates this process. In the present study, we investigated the role of A-kinase anchoring protein 3 (AKAP3) during mouse spermiogenesis, using both mouse genetics and proteomics. It was found that AKAP3 is essential for the formation of the specific subcellular structure of the sperm flagellum, motility of sperm and male fertility. Additionally, lack of AKAP3 caused global changes of the sperm proteome and mislocalization of sperm proteins, including accumulation of RNA metabolism and translation factors and displacement of PKA subunits in mature sperm, which may underlie misregulated PKA activity and immotility in sperm. Interestingly, sperm lacking a complete fibrous sheath from both Akap3 and Akap4 null mice accumulated F-actin filaments and morphological defects during post-testicular maturation in the epididymis. These results suggest that the subcellular structures of sperm could be formed via independent pathways, and elucidate the roles of AKAP3 during the coordinated synthesis and organization of the sperm proteome and sperm morphology. [ABSTRACT FROM AUTHOR]

Purpose: Children may require the long‐term use of a central venous catheter (CVC) for treatment. Multiple revisions may be required when these catheters malfunction. We herein reported our experience of utilizing the fibrous sheath in tunnelled CVCs for the revision. Methods: A retrospective review was conducted from July 2008 to June 2018 on all children with the revision of tunnelled CVCs using the fibrous sheath. Results: Sixteen children underwent 31 revisions of a tunnelled CVC (16 Broviac/Hickman catheters, 15 Permcath for haemodialysis). The age ranged from 6 months to 16 years old (mean 3.8 years old). The indications for CVC revision included blockage (n = 14), breakage (n = 9) and dislodgement (n = 6). The mean time interval between CVC insertion and revision was 10.1 months (range: 20 days to 39 months). The fibrous sheath could be used for re‐insertion in 27 revisions (success rate: 87%). The failure was associated with the upsizing of the CVC (P =.003). In three of the four failed cases, a new CVC could be inserted by the cut‐down technique through the same venous access site. The overall success rate of CVC revision using the same venous access was 97%. There was no 30‐day postoperative infective complication. Conclusions: The fibrous sheath of tunnelled CVCs is a reliable access for CVC re‐insertion. This helps to preserve the same venous access site for repeated revision of CVCs. This is extremely important in those children who depend on long‐term CVC usage. [ABSTRACT FROM AUTHOR]

Sperm fibrous sheath, a unique cytoskeletal structure, is implicated in various sperm physiological functions, such as sperm maturation, motility and capacitation. AKAP4 has been described to be required for structural and functional integrity of the fibrous sheath. We generated Akap4 -knockout mice line using CRISPR-Cas9 system. Cytomorphology and motility of sperm and testes were studied, confirming loss of Akap4 led to abnormal sperm morphology, motility and infertility. The proteomic components of testes were studied and Akap4 was found to be significantly decreased in the Akap4 -knockout mice. Testis single-cell RNA sequencing and analysis revealed three genes with significant change in the general cell population, i.e., Akap4 , Haspin , and Ccdc38. The single-cell RNA expression profiles also showed that the major difference between Akap4 -knockout and wild-type testes existed in the elongating cell cluster, where in the Akap4 -knockout testes, a subgroup of elongating cells with marker genes involved in cell adhesion and migration were increased, while a subgroup of elongating cells marked by mitochondrial sheath genes were decreased. Our results revealed the complex and well-coordinated procedures of spermatogenesis, and substantiated Akap4 's indispensable roles in the integrity of sperm flagellum and the step-wise maturation of spermatozoa. [ABSTRACT FROM AUTHOR]

Motile flagella are crucial for human fertility and embryonic development. The distal tip of the flagellum is where growth and intra‐flagellar transport are coordinated. In most model organisms, but not all, the distal tip includes a 'singlet region', where axonemal doublet microtubules (dMT) terminate and only complete A‐tubules extend as singlet microtubules (sMT) to the tip. How a human flagellar tip is structured is unknown. Here, the flagellar tip structure of human spermatozoa was investigated by cryo‐electron tomography, revealing the formation of a complete sMT from both the A‐tubule and B‐tubule of dMTs. This different tip arrangement in human spermatozoa shows the need to investigate human flagella directly in order to understand their role in health and disease. [ABSTRACT FROM AUTHOR]

Study Question: Can whole-exome sequencing (WES) of infertile patients identify new genes responsible for multiple morphological abnormalities of the sperm flagella (MMAF)?Summary Answer: WES analysis of 78 infertile men with a MMAF phenotype permitted the identification of four homozygous mutations in the fibrous sheath (FS) interacting protein 2 (FSIP2) gene in four unrelated individuals.What Is Known Already: The use of high-throughput sequencing techniques revealed that mutations in the dynein axonemal heavy chain 1 (DNAH1) gene, and in the cilia and flagella associated protein 43 (CFAP43) and 44 (CFAP44) genes account for approximately one-third of MMAF cases thus indicating that other relevant genes await identification.Study Design, Size, Duration: This was a retrospective genetics study of 78 patients presenting a MMAF phenotype who were recruited in three fertility clinics between 2008 and 2015. Control sperm samples were obtained from normospermic donors. Allelic frequency for control subjects was derived from large public databases.Participants/materials, Setting, Methods: WES was performed for all 78 subjects. All identified variants were confirmed by Sanger sequencing. Relative mRNA expression levels for the selected candidate gene (FSIP2) was assessed by quantitative RT-PCR in a panel of normal human and mouse tissues. To characterize the structural and ultrastructural anomalies present in patients' sperm, immunofluorescence (IF) was performed on sperm samples from two subjects with a mutation and one control and transmission electron microscopy (TEM) analyses was performed on sperm samples from one subject with a mutation and one control.Main Results and the Role Of Chance: We identified four unrelated patients (4/78, 5.1%) with homozygous loss of function mutations in the FSIP2 gene, which encodes a protein of the sperm FS and is specifically expressed in human and mouse testis. None of these mutations were reported in control sequence databases. TEM analyses showed a complete disorganization of the FS associated with axonemal defects. IF analyses confirmed that the central-pair microtubules and the inner and outer dynein arms of the axoneme were abnormal in all four patients carrying FSIP2 mutations. Importantly, and in contrast to what was observed in patients with MMAF and mutations in other MMAF-related genes (DNAH1, CFAP43 and CFAP44), mutations in FSIP2 led to the absence of A-kinase anchoring protein 4 (AKAP4).Limitations, Reasons For Caution: The low number of biological samples and the absence of a reliable anti-FSIP2 antibody prevented the formal demonstration that the FSIP2 protein was absent in sperm from subjects with a FSIP2 mutation.Wider Implications Of the Findings: Our findings indicate that FSIP2 is one of the main genes involved in MMAF syndrome. In humans, genes previously associated with a MMAF phenotype encoded axonemal-associated proteins (DNAH1, CFAP43 and CFAP44). We show here that FSIP2, a protein of the sperm FS, is also logically associated with MMAF syndrome as we showed that it is necessary for FS assembly and for the overall axonemal and flagellar biogenesis. As was suggested before in mouse and man, our results also suggest that defects in AKAP4, one of the main proteins interacting with FSIP2, would induce a MMAF phenotype. Finally, this work reinforces the demonstration that WES sequencing is a good strategy to reach a genetic diagnosis for patients with severe male infertility phenotypes.Study Funding/competing Interest(s): This work was supported by the following grants: the 'MAS-Flagella' project financed by the French ANR and the DGOS for the program PRTS 2014 (14-CE15) and the 'Whole genome sequencing of patients with Flagellar Growth Defects (FGD)' project financed by the Fondation Maladies Rares for the program Séquençage à haut débit 2012. The authors have no conflict of interest. [ABSTRACT FROM AUTHOR]

Ca2+-binding tyrosine-phosphorylation-regulated protein (CABYR) has been implicated in sperm physiological function in several in vitro studies. It has also been implicated as a potential cause of and diagnostic tool in asthenozoospermic human males. CABYR is known to be localized to the fibrous sheath, an accessory structure in the flagellar principal piece. Utilizing the CRISPR--Cas9 technology, we have knocked out this gene inmice to understand its role inmale fertility. Cabyr-knockout male mice showed severe subfertility with a defect in sperm motility as well as a significant disorganization in the fibrous sheath. Further, abnormal configuration of doublet microtubules was observed in the Cabyr-knockout spermatozoa, suggesting that the fibrous sheath is important for the correct organization of the axoneme. Our results show that it is the role of CABYR in the formation of the fibrous sheath that is essential for male fertility. [ABSTRACT FROM AUTHOR]

Abstract: Genes exclusively expressed in male germ cells encode proteins that play important roles in spermatogenesis and fertilization. In this study, we investigated the expression of a novel spermatogenic cell-specific gene known as amyotrophic lateral sclerosis 2 chromosome region candidate 12 (Als2cr12). Our in silico and in vitro analyses revealed that the mouse Als2cr12 gene produces two transcript isoforms by alternative splicing and that one of the isoforms is unique to spermatogenic cells. Using an antibody against the ALS2CR12 protein, we found that a protein from the germ cell-specific Als2cr12 transcript is present in mature sperm from the epididymis as well as germ cells in the testis. Further analysis of the ALS2CR12 protein in sperm disclosed the localization of the protein in the sperm tail. Specifically, our data suggest that the ALS2CR12 protein is associated with the fibrous sheath in the sperm flagellum. Thus, our study provides the first information regarding the expression of the Als2cr12 gene at the transcriptional, protein and cellular levels. [Copyright &y& Elsevier]

Abstract: We report two cases of encapsulated indwelling central venous catheter in adults. The patients involved were treated by chemotherapy and their catheter had been implanted for 15 and 6 years. However, this complication is well-known in children and a similar problem is frequently observed with pacemaker leads removal. The difficulty to remove the device is due to fibrous encapsulation of indwelling catheters in the vein wall. In all cases reported, the catheter was left in place. The potential main complications not yet observed are infection of the catheter and venous thrombosis. [Copyright &y& Elsevier]

Glyceraldehyde-3-phosphate dehydrogenase (GAPDs) was purified from human sperms and properties of the enzyme were investigated. After sonication of sperms, the most part of GAPDs is associated with the insoluble cell fraction. Trypsin treatment results in the cleavage of part of the N-terminal domain of the enzyme yielding a soluble fragment that was purified by hydrophobic chromatography on Phenyl-Sepharose. The isolated fragment was shown to be a tetramer with molecular weight of approximately 150 kD (according to Blue Native PAGE) and composed of subunits of 40 kD (according to SDS-PAGE). The specific activity of the isolated fragment reached 374 U/mg. It is supposed that GAPDs exists in sperms as the tetrameric molecule bound to the fibrous sheath of the flagellum through the N-terminus of one or two subunits. Comparative study of the amino acid sequences of mammalian GAPDs revealed conservative cysteine residues (C21, C94, and C150) that are specific for the sperm isoenzyme and absent in the somatic isoenzyme. Residue C21 can be involved in the formation of the disulfide bond between the N-terminal domain of GAPDs and fibrous sheath proteins. [ABSTRACT FROM AUTHOR]

Abstract: Enzymes in the glycolytic pathway of mammalian sperm are modified extensively and are localized in the flagellum, where several are tightly bound to the fibrous sheath. This study provides the first evidence for three novel aldolase isozymes in mouse sperm, two encoded by Aldoart1 and Aldoart2 retrogenes on different chromosomes and another by Aldoa_v2, a splice variant of Aldoa. Phylogenetic analyses and comparative genomics indicate that the retrogenes and splice variant have remained functional and have been under positive selection for millions of years. Their expression is restricted to the male germline and is tightly regulated at both transcriptional and translational levels. All three isozymes are present only in spermatids and sperm and have distinctive features that may be important for localization in the flagellum and/or altered metabolic regulation. Both ALDOART1 and ALDOA_V2 have unusual N-terminal extensions not found in other aldolases. The N-terminal extension of ALDOA_V2 is highly conserved in mammals, suggesting a conserved function in sperm. We hypothesize that the N-terminal extensions are responsible for localizing components of the glycolytic pathway to the fibrous sheath and that this localization is required to provide sufficient ATP along the length of the flagellum to support sperm motility. [Copyright &y& Elsevier]

Abstract: The longest part of the sperm flagellum, the principal piece, contains the fibrous sheath, a cytoskeletal element unique to spermiogenesis. We performed mass spectrometry proteomics on isolated human fibrous sheaths identifying a unique ADP/ATP carrier protein, SFEC [AAC4], seven glycolytic enzymes previously unreported in the human sperm fibrous sheath, and sorbitol dehydrogenase. SFEC, pyruvate kinase and aldolase were co-localized by immunofluorescence to the principal piece. A homology model constructed for SFEC predicted unique residues at the entrance to the nucleotide binding pocket of SFEC that are absent in other human ADP/ATP carriers, suggesting opportunities for selective drug targeting. This study provides the first evidence of a role for an ADP/ATP carrier family member in glycolysis. The co-localization of SFEC and glycolytic enzymes in the fibrous sheath supports a growing literature that the principal piece of the flagellum is capable of generating and regulating ATP independently from mitochondrial oxidation in the mid-piece. A model is proposed that the fibrous sheath represents a highly ordered complex, analogous to the electron transport chain, in which adjacent enzymes in the glycolytic pathway are assembled to permit efficient flux of energy substrates and products with SFEC serving to mediate energy generating and energy consuming processes in the distal flagellum, possibly as a nucleotide shuttle between flagellar glycolysis, protein phosphorylation and mechanisms of motility. [Copyright &y& Elsevier]

Morphogenesis of the mammalian sperm flagellum is characterized by the assembly of axonemal and peri-axonemal structures. The incorporation of mitochondria into the flagellum results from complex cellular events, including flagellum compartmentalization and membrane and organelle reorganization. These events are striking in the annulus, which progressively relocates from the neck to the principal piece of the flagellum. This study presents a human sperm phenotype with failure of the annulus relocation, absence of mitochondrial sheath and a fibrous sheath at intermediate step of assembly. The sperm nucleus was fully condensed but with deep invaginations engulfing the acrosome. The distal pole of some mitochondria exhibited an unusual dense substance. This rare human sperm phenotype was found in a consanguineous patient, suggesting a genetic origin. These anomalies raise the question of the mechanisms that lead to impairment of both the annulus relocation and the deposit of proteins on the fibrous sheath during spermiogenesis. [ABSTRACT FROM AUTHOR]