Secretion and degradation of parathormone as a function of intracellular maturation of hormone pools

The biosynthesis, processing, and secretion of parthormone and the effect of calcium on these processes were measured in dispersed porcine parthyroid cells incubated with [(35)S]methionine. Proparathormone was detected at 10 min, the earliest time measured, and was rapidly and apparently quantitatively converted to parathormone. The half-life of the prohomormone pool was 15 min. Secretion of parathormone was detected by 20 min. In pulse-chase experiments there was a period between 20 and 40 min during which the wave of newly-synthesized parathormone was secreted. After 40 min during little additional radioactive hormone was secreted, but dibutyryl cyclic AMP, an agent that can mobilize stored parathormone, when added to the incubation mixtures enhanced radioactive parathormone secretion but only after 60 min, although it increased net hormone secretion as determined by radioimmunoassay to the same extent at all times studied. When the ionized calcium concentration of the medium was lowered, more radioactive hormone was secreted at all times but the effect was greatest on that hormone that was synthesized less than 60 min previously ; however, net hormone secretion in contrast to radioactive hormone was enhanced equally at all intervals. These data could mean that the refractoriness to secretion of parathormone 40-60 min of age was related to maturation of secretory container preparatory to storage. Low calcium (0.5 mM) stimulated hormone secretion up to fivefold compared to high calcium (3.0 mM) but did not affect synthesis of parathormone or proparathormne or conversion of the latter to hormone. During processing at least 70 percent of the intracellular parathormone was lost, presumably through proteolysis and this degradation was greater at high calcium. These data have been interpreted in light of the concept that two secretable pools of parathormone exist within the parathyroid.     

Plasma levels of progesterone, provera, oestradiol-17beta, and 13,14-dihydro-15-keto-prostaglandin F in cows treated with provera-impregnated intravaginal sponges.

Plasma concentrations of progesterone and Provera were measured daily in 3 cows during 21 days of treatment with Provera-impregnated intravaginal sponges. Plasma concentrations of oestradiol-17beta and 13,14-dihydro-15-keto-prostaglandin F (PGFM) were measured hourly from 5 h before until 62 h after sponge removal. The profile of progesterone concentrations indicated that luteolysis occurred at the expected time (Days 19 to 23 of the cycle), even though plasma Provera concentrations were 150-250 pg/ml. The occurrence of peaks of PGFM after sponge withdrawal suggests that PGF-2alpha release is stimulated by falling levels of progestagen.     

Effect of Hailey-Hailey Disease mutations on the function of a new variant of human secretory pathway Ca2+/Mn2+-ATPase (hSPCA1)

ATP2C1, encoding the human secretory pathway Ca2+/Mn2+ ATPase (hSPCA1), was recently identified as the defective gene in Hailey-Hailey Disease (HHD), an autosomal dominant skin disorder characterized by persistent blisters and erosions. To investigate the underlying cause of HHD, we have analyzed the changes in expression level and function of hSPCA1 caused by mutations found in HHD patients. Mutations were introduced into hSPCA1d, a novel splice variant expressed in keratinocytes, described here for the first time. Encoded by the full-length of optional exons 27 and 28, hSPCA1d was longer than previously identified splice variants. The protein competitively transported Ca2+ and Mn2+ with equally high affinity into the Golgi of COS-1 cells. Ca2+- and Mn2+-dependent phosphoenzyme intermediate formation in forward (ATP-fuelled) and reverse (Pi-fuelled) directions was also demonstrated. HHD mutant proteins L341P, C344Y, C411R, T570I, and G789R showed low levels of expression, despite normal levels of mRNA and correct targeting to the Golgi, suggesting instability or abnormal folding of the mutated hSPCA1 polypeptides. P201L had little effect on the enzymatic cycle, whereas I580V caused a block in the E1 approximately P --> E2-P conformational transition. D742Y and G309C were devoid of Ca2+- and Mn2+-dependent phosphoenzyme formation from ATP. The capacity to phosphorylate from Pi was retained in these mutants but with a loss of sensitivity to both Ca2+ and Mn2+ in D742Y and a preferential loss of sensitivity to Mn2+ in G309C. These results highlight the crucial role played by Asp-742 in the architecture of the hSPCA1 ion-binding site and reveal a role for Gly-309 in Mn2+ transport selectivity.     

Do the maximum sizes,ages and patterns of growth of three reef‐dwelling labrid species at two latitudes differ in a manner conforming to the metabolic theory of ecology?

The size and age data and patterns of growth of three abundant, reef‐dwelling and protogynous labrid species (Coris auricularis, Notolabrus parilus and Ophthalmolepis lineolata) in waters off Perth at c. 32° S and in the warmer waters of the Jurien Bay Marine Park (JBMP) at c. 30° S on the lower west coast of Australia are compared. Using data for the top 10% of values and a randomization procedure, the maximum total length (L_T) and mass of each species and the maximum age of the first two species were estimated to be significantly greater off Perth than in the JBMP (all P < 0·001) and the maximum ages of O. lineolata in the two localities did not differ significantly (P > 0·05). These latitudinal trends, thus, typically conform to those frequently exhibited by fish species and the predictions of the metabolic theory of ecology (MTE). While, in terms of mass, the instantaneous growth rates of each species were similar at both latitudes during early life, they were greater at the higher latitude throughout the remainder and thus much of life, which is broadly consistent with the MTE. When expressed in terms of L_T, however, instantaneous growth rates did not exhibit consistent latitudinal trends across all three species. The above trends with mass, together with those for reproductive variables, demonstrate that a greater amount of energy is directed into somatic growth and gonadal development by each of these species at the higher latitude. The consistency of the direction of the latitudinal trends for maximum body size and age and pattern of growth across all three species implies that each species is responding in a similar manner to differences between the environmental characteristics, such as temperature, at those two latitudes. The individual maximum L_T, mass and age and pattern of growth of O. lineolata at a higher and thus cooler latitude on the eastern Australian coast are consistent with the latitudinal trends exhibited by those characteristics for this species in the two western Australian localities. The implications of using mass rather than length as the indicator variable when comparing the maximum sizes of the three species and the trends exhibited by the instantaneous growth rates of those species at different latitudes are explored. Although growth curves fitted to both the L_T and masses at age for the males of each species lay above those for their females, this would not have influenced the conclusions drawn from common curves for both sexes.     

Highly branched poly(N-isopropylacrylamide) for use in protein purification

Poly(N-isopropylacrylamide)s with imidazole endgroups were used to separate a histidine-tagged protein fragment directly from a crude cell lysate. The polymers display a lower critical solution temperature that can be tuned to occur at a range of subambient temperatures. UV-visible spectra indicated differences in the binding in aqueous media of Cu(II) and Ni(II) to the imidazole endgroups. These changes in the UV-visible spectra were reflected in the solution/aggregation behavior of the polymers as studied by dynamic light scattering. The addition of Cu(II) disaggregated the polymers, and the polymer coil swelled. On the other hand, when Ni(II) was added the polymers remained aggregated in aqueous media. The polymers were used to purify residues 230-534 of the histidine-tagged breast cancer susceptibility protein his6-BRCA1. Cu(II) was found to be better suited to the formation of useful polymer-metal ion-protein complexes that display cloud points, since Ni(II)/polymer mixtures generated very little purified protein. The polymers were synthesized using a previously reported variation of the reversible addition-fragmentation chain termination (RAFT) methodology, using the chain transfer agent 3H-imidazole-4-carbodithioic acid 4-vinyl benzyl ester with N-isopropylacrylamide (NIPAM).     

Age,growth and reproductive life-history characteristics infer a high population productivity for the sustainably fished protogynous hermaphroditic yellowspotted rockcod (Epinephelus areolatus) in north-western Australia

The yellowspotted rockcod, Epinephelus areolatus, is a small-sized grouper that is widely distributed throughout the Indo-Pacific, where it forms a valuable component of the harvest derived from multispecies fisheries along continental and insular shelves. Samples of E. areolatus were collected from 2012 to 2018 from commercial catches and research surveys in the Kimberley, Pilbara and Gascoyne regions of north-western Australia to improve the understanding of the life history, inherent vulnerability and stock status of this species. Histological analysis of gonads (n = 1889) determined that E. areolatus was a monandric protogynous hermaphrodite. Non-functional spermatogenic crypts were dispersed within the ovaries of 23% of mature functioning females; nonetheless, these crypts were not observed during the immature female phase. The length and age at which 50% of females matured were 266 mm total length (L_T) and 2.7 years, respectively. The spawning period was protracted over 10–12 months of the year with biannual peaks at the start of spring and autumn (i.e., September and March) when the photoperiod was at its mid-range (i.e., 12.1 h). Estimates of the lengths and ages at which 50% of E. areolatus change sex from female to male were very similar (i.e., <5% difference) between the Kimberley and Pilbara regions, i.e., of 364 and 349 mm L_T and of 7.9 and 7.3 years, respectively. A maximum age of 19 years was observed in all three regions, but there was significant regional variation in growth. These variations in growth were not correlated with latitude; instead a parabolic relationship was evident, where the smallest mean length-at-age and fastest growth rates (k) occurred in the mid-latitudes of the Pilbara region. In the Kimberley and Pilbara regions, individuals were not fully selected by commercial fish traps until 5–6 years of age, hence, several years after reaching maturity. These life-history characteristics infer a high population productivity, which underpins the sustainable harvest of this species, despite comprising the largest catches of all epinephelids in the multispecies tropical fisheries across north-western Australia.     

Effect of systemic or intrauterine injections of indomethacin on plasma oxytocin-neurophysin concentrations in ewes over the time of normal corpus luteum regression

This study was undertaken to investigate the effect of systemic or intrauterine injections of indomethacin, a known prostaglandin (PG) synthetase inhibitor, on peripheral plasma oxytocin-associated neurophysin (OT-N) concentrations in ewes over the time of expected luteolysis. In the first experiment, 9 ewes were given i.m. injections of indomethacin (4 mg/kg live weight, n = 4) or vehicle (n = 5) 3 times/day over Days 13-15 of the estrous cycle. Blood samples were collected at hourly intervals from 0700 h on Day 13 to 1800 h on Day 15 post-estrus. In the second experiment, indomethacin (20 mg, n = 5) or the injection vehicle (n = 4) was given twice daily into the uterine horn over Days 12-14 post-estrus. Blood samples were collected at hourly intervals from Day 12 to 14. In the third experiment, 4 additional ewes were bled at 5-min intervals from 1200 to 1600 h on Day 13 of the estrous cycle. Plasma samples were analyzed for OT-N and 13,14-dihydro-15-keto-prostaglandin F2 alpha (PGFM) to provide an indirect index for ovarian oxytocin and uterine prostaglandin F2 alpha release, respectively. Results from the first experiment indicated that surges in plasma OT-N concentrations occurred in the vehicle-treated ewes but were suppressed in ewes given systemic injections of indomethacin. Intrauterine indomethacin injections did not cause a significant reduction in the maximum peak height or number of peaks when compared with the control ewes. In the third experiment, there was a marked increase in plasma OT-N concentrations, but no significant rise in plasma PGFM concentrations in one ewe.(ABSTRACT TRUNCATED AT 250 WORDS)