Russell C. Jones

In the platypus a meiotic chain of ten sex chromosomes shares genes with the bird Z and mammal X chromosomes

Two centuries after the duck-billed platypus was discovered, monotreme chromosome systems remain deeply puzzling. Karyotypes of males, or of both sexes, were claimed to contain several unpaired chromosomes (including the X chromosome) that form a multi-chromosomal chain at meiosis. Such meiotic chains exist in plants and insects but are rare in vertebrates. How the platypus chromosome system works to determine sex and produce balanced gametes has been controversial for decades. Here we demonstrate that platypus have five male-specific chromosomes (Y chromosomes) and five chromosomes present in one copy in males and two copies in females (X chromosomes). These ten chromosomes form a multivalent chain at male meiosis, adopting an alternating pattern to segregate into XXXXX-bearing and YYYYY-bearing sperm. Which, if any, of these sex chromosomes bears one or more sex-determining genes remains unknown. The largest X chromosome, with homology to the human X chromosome, lies at one end of the chain, and a chromosome with homology to the bird Z chromosome lies near the other end. This suggests an evolutionary link between mammal and bird sex chromosome systems, which were previously thought to have evolved independently.

Micropuncture and cannulation studies of fluid composition and transport in the ductuli efferentes testis of the rat: comparisons with the homologous metanephric proximal tubule

Luminal fluids were collected in vivo by micropuncture and cannulation from the rete testis, efferent ducts and ductus epididymidis of the rat to determine the composition of efferent duct fluids and the rates of reabsorption of water and solutes by the efferent ducts. The concentration of spermatozoa increased by a factor of about 25 from 2.42 x 10(4) microliters‐1 in the fluid from the rete testis to 6.00 x 10(5) microliters‐1 in fluid at the end of the efferent ducts, indicating that 96.2% of the fluid leaving the testis is reabsorbed from the lumen of the efferent ducts. Most of this reabsorption (70.9% or 33.4 microliters h‐1) occurs in the region between the rete testis and the middle of the coni vasculosi, with only 25.1% (11.8 microliters h‐1) occurring between the coni and the beginning of the ductus epididymidis. However, reabsorption across the epithelium occurs at about the same rate in both regions, with the proximal region reabsorbing 17.2 microliters cm‐2 h‐1 (70.9% of fluid entering the region) and the distal region reabsorbing 12.2 microliters cm‐2 h‐1 (86.1% of fluid entering the region). Consequently, the fluid reabsorption rate for the whole efferent duct system (15.6 microliters cm‐2 h‐1) is similar to the values for individual regions. The principal solutes in luminal fluids from the efferent ducts are Na+ (137‐144 mM) and Cl‐ (113‐130 mM). The estimated sum contribution of Na+, Cl‐ and K+ to the osmotic pressure of luminal fluids was approximately 80% at each site sampled in the efferent ducts. The osmotic pressure of luminal fluid samples (301‐307 mosmol kg‐1) did not vary significantly along the ducts or differ significantly from that of blood plasma. The results demonstrate that there is a net reabsorption in the efferent ducts of nearly all the testicular output of water and inorganic electrolytes, and most of the protein, and that, in comparison, the ductus epididymidis is a negligible site of net fluid reabsorption. The results indicate that the ductus epididymidis, rather than the efferent ducts, is the site of accumulation of high concentrations of specific organic compounds like inositol. The efferent ducts are similar to the homologous proximal tubules of the metanephric kidney in that the luminal electrolyte composition (principal solutes Na+ and Cl‐) and osmotic pressure remain relatively stable and that fluid reabsorption is close to isotonic and occurs at the same rate as the reabsorption of Na+.

Tyrosine Phosphorylation of HSP-90 During Mammalian Sperm Capacitation

Abstract The process of sperm capacitation is correlated with activation of a signal transduction pathway leading to protein tyrosine phosphorylation. Whereas phosphotyrosine expression is an essential prerequisite for fertilization, the proteins that are phosphorylated during capacitation have not yet been identified. In the present study, we observed that a major target of this signaling pathway is the molecular chaperone protein, heat shock protein (HSP)-86, a member of the HSP-90 family of HSPs. We used cross-immunoprecipitation experiments to confirm the tyrosine phosphorylation of HSP-86, a process that is not inhibited by the ansamycin antibiotic, geldanamycin. The general significance of these findings was confirmed by studies in which HSP-90 was also found to be tyrosine phosphorylated in human and rat spermatozoa when incubated under conditions that support capacitation. To our knowledge, these results represent the first report of a protein that undergoes tyrosine phosphorylation during mouse sperm capacitation and the first study implicating molecular chaperones in the processes by which mammalian spermatozoa gain the ability to fertilize the oocyte.

Mammalian epididymal proteome.

In all mammalian species, the final differentiation of the male germ cell occurs in the epididymal duct where the spermatozoa develop the ability to be motile and fertilize an ovum. Understanding of these biological processes is the key to understanding and controlling male fertility. Comparative studies between several mammals could be an informative approach to finding common sperm modifications which are not species-specific. The new global biological approaches such as transcriptomes and proteomes provide considerable information which can be used for such comparative approaches. This report summarizes our proteomic studies of the epididymis of several mammals, including humans.

Endogenous Redox Activity in Mouse Spermatozoa and Its Role in Regulating the Tyrosine Phosphorylation Events Associated with Sperm Capacitation

Abstract We investigated the role of endogenous redox activity in regulating the signal transduction pathway leading to tyrosine phosphorylation in mouse spermatozoa. Endogenous redox activity was monitored using a luminol-peroxidase chemiluminescent probe. Chemiluminescence increased in spermatozoa that were actively undergoing cAMP-mediated tyrosine phosphorylation events associated with capacitation and was inhibited in a dose-dependent manner by addition of catalase or diphenylene iodonium, both of which also inhibited tyrosine phosphorylation within the cell at points downstream of cAMP. Excluding bicarbonate from the incubation medium reduced the redox activity of sperm by 80–90% and dramatically reduced tyrosine phosphorylation. This study provides the first evidence that tyrosine phosphorylation associated with capacitation in mouse spermatozoa is redox regulated by a flavinoid-containing enzyme involving mediation by hydrogen peroxide. Bicarbonate regulated the redox activity of mouse spermatozoa, and this regulation may contribute to the impact of this anion on tyrosine phosphorylation during capacitation of mouse spermatozoa.

The role of Na+-H+ exchange in fluid and solute transport in the rat efferent ducts.

In vivo microperfusion techniques were used to investigate the role of Na+‐H+ exchange in the efferent ducts of the rat. Individual efferent ducts were perfused with a Krebs‐Ringer bicarbonate solution (KRB) containing 0, 1, 3, 5 or 7·5 mM amiloride. Concentrations of 1‐5 mM amiloride inhibited fluid reabsorption from the efferent ducts in a linear dose‐dependent manner with an apparent Km of 3 mM. Inhibition was maximal at 5 mM with reabsorption reduced by about 70%. The effects of amiloride were completely reversible and there was little effect of amiloride on luminal osmolality and concentrations of Na+, Cl− or K+. It is concluded that Na+‐H+ exchange is one of the principal mechanisms responsible for fluid and electrolyte reabsorption in the efferent ducts and offers a means by which the efferent ducts are able to achieve flow‐dependent, autoregulated fluid reabsorption.

Renewal and proliferation of spermatogonia during spermatogenesis in the Japanese quail, Coturnix coturnix japonica

SummaryFour different types of spermatogonia were identified in the seminiferous tubules of the Japanese quail: a dark type A (Ad), 2 pale A type (Ap1 and Ap2), and a type B. A model is proposed describing the process of spermatogonial development in the quail. The Ad spermatogonia are considered to be the stem cells. Each divides to produce a new Ad spermatogonium and a Ap1 spermatogonium during Stage IX of the cycle of the seminiferous epithelium. An Ap1 spermatogonium produces two Ap2 spermatogonia during Stage II of the cycle, Ap2 spermatogonia produce four type B spermatogonia during Stage VI of the cycle, and type B spermatogonia produce eight primary spermatocytes during Stage III of the cycle. Consequently, 32 spermatids can result from each division of an Ad spermatogonium. Spermatogonial development in the quail differs from the process described in mammals in that there are fewer mitotic divisions and they are all synchronized with the cycle of the seminiferous epithelium. It is suggested that the fewer mitotic divisions explain why a smaller area of the seminiferous tubule is occupied by a cellular association in the quail than in mammals like the rat, ram and bull. The duration of spermatogenesis from the division of the Ad spermatogonia to sperm release from the seminiferous epithelium was estimated to be 12.77 days.

Transport of IgG across the Blood-Luminal Barrier of the Male Reproductive Tract of the Rat and the Effect of Estradiol Administration on Reabsorption of Fluid and IgG by the Epididymal Ducts

Abstract In rats immunized systemically with tetanus toxoid the concentration of specific anti-tetanus-toxoid-specific IgG in fluid from the rete testis and cauda epididymidis were respectively 0.6% and 1.4% the concentration in blood serum. The extratesticular duct system reabsorbed 97% of the IgG and 99% of the fluid leaving the rete, but estradiol administration affected the site of reabsorption. In untreated rats, the ductuli efferentes reabsorbed 94% of the IgG and 96% of the fluid leaving the rete, whereas estradiol-treated rats reabsorbed 83% of the IgG and 86% of the fluid, and the ductus epididymidis fully compensated for these different effects of estradiol on the ductuli efferentes. The concentrations of IgG in secretions of the seminal vesicles and prostate gland were lower (0.1% and 0.3% respectively of the titers in blood serum) than in fluids from the extratesticular ducts, and were not affected by the administration of estradiol. RT-PCR showed that Fcgrt (neonatal Fc receptor, also known as FcRn) is expressed in the reproductive ducts, where IgG is probably transported across epithelium, being particularly strong in the ductuli efferentes (where most IgG was reabsorbed) and distal caput epididymidis. It is concluded that IgG enters the rete testis and is concentrated only 2.5-fold along the extratesticular duct system, unlike spermatozoa, which are concentrated 95-fold. Further, the ductus epididymidis can recognize and compensate for changes in function of the ductuli efferentes.

Restricted Entry of IgG into Male and Female Rabbit Reproductive Ducts Following Immunization with Recombinant Rabbit PH-20

PROBLEM: Successful immunocontraception using sperm antigens is dependent on achieving sufficient sperm‐specific antibody in the reproductive ducts to prevent fertilization. The blood : luminal barrier of the male and female reproductive ducts must be overcome for this to occur. We have, therefore, investigated the relative titers of antigen‐specific immunoglobulin G (IgG) in luminal fluids collected from male and female rabbit reproductive ducts following immunization with recombinant rabbit PH‐20 (rPH‐20).

Spermiogenesis and spermiation in a monotreme mammal, the platypus, Ornithorhynchus anatinus.

Spermatogenesis in the platypus (Ornithorhynchus anatinus) is of considerable biological interest as the structure of its gametes more closely resemble that of reptiles and birds than marsupial or eutherian mammals. The ultrastructure of 16 steps of spermatid development is described and provides a basis for determining the kinetics of spermatogenesis. Steps 1–3 correspond to the Golgi phase of spermatid development, steps 4–8 correspond to the cap phase, steps 9–12 are the acrosomal phase, and steps 13–16 are the maturation phase. Acrosomal development follows the reptilian model and no acrosomal granule is formed. Most other features of spermiogenesis are similar to processes in reptiles and birds. However, some are unique to mammals. For example, a thin, lateral margin of the acrosome of platypus sperm expands over the nucleus as in other mammals, and more than in reptiles and birds. Also, a tubulobulbar complex develops around the spermatid head, a feature which appears to be unique to mammals. Further, during spermiation the residual body is released from the caudal end of the nucleus of platypus sperm leaving a cytoplasmic droplet located at the proximal end of the middle piece as in marsupial and eutherian mammals. Other features of spermiogenesis in platypus appear to be unique to monotremes. For example, nuclear condensation involves the formation of a layer of chromatin granules under the nucleolemma, and development of the fibrous sheath of the principal piece starts much later in the platypus than in birds or eutherian mammals.