STIM1 is a MT-plus-end-tracking protein involved in remodeling of the ER

Stromal interaction molecule 1 (STIM1) is a transmembrane protein that is essential for store-operated Ca(2+) entry, a process of extracellular Ca(2+) influx in response to the depletion of Ca(2+) stores in the endoplasmic reticulum (ER) (reviewed in [1-4]). STIM1 localizes predominantly to the ER; upon Ca(2+) release from the ER, STIM1 translocates to the ER-plasma membrane junctions and activates Ca(2+) channels (reviewed in [1-4]). Here, we show that STIM1 directly binds to the microtubule-plus-end-tracking protein EB1 and forms EB1-dependent comet-like accumulations at the sites where polymerizing microtubule ends come in contact with the ER network. Therefore, the previously observed tubulovesicular motility of GFP-STIM1 [5] is not a motor-based movement but a traveling wave of diffusion-dependent STIM1 concentration in the ER membrane. STIM1 overexpression strongly stimulates ER extension occurring through the microtubule "tip attachment complex" (TAC) mechanism [6, 7], a process whereby an ER tubule attaches to and elongates together with the EB1-positive end of a growing microtubule. Depletion of STIM1 and EB1 decreases TAC-dependent ER protrusion, indicating that microtubule growth-dependent concentration of STIM1 in the ER membrane plays a role in ER remodeling.     

Electrogenic Cl<Superscript>−</Superscript> secretion does not occur in the ileum of the Australian common brushtail possum, <Emphasis Type="Italic">Trichosurus vulpecula</Emphasis>, due to low levels of expression of the NaK2Cl cotransporter,NKCC1

The colon of the brushtail possum does not have an electrogenic secretory response. Given the functional significance of electrogenic Cl⁻ secretion in the intestine of eutherian mammals, we have investigated the secretory response in the small intestine of this marsupial. In the Ussing chamber cAMP-dependent secretagogues stimulated a sustained increase in ileal short-circuit current (Isc), whereas Ca²⁺-dependent secretagogues induced a transient increase. Both the responses were inhibited by mucosal addition of the anion channel blocker 5-nitro-2-(3-phenylpropylamino) benzoic acid (100 μmol l⁻¹), consistent with an anion secretory response. However, the responses were not inhibited by serosal bumetanide (10 μmol l⁻¹) and were independent of bath Cl⁻, indicating that the stimulated ileal Isc does not involve electrogenic Cl⁻ secretion driven by the NaK2Cl cotransporter, NKCC1. Consistent with this, there were low levels of NKCC1 expression in the ileal epithelium. In particular, NKCC1 expression in the ileal crypt cells was comparable to that of the villous cells. This differs from eutherian mammals where high levels of NKCC1 expression in the ileal crypt cells are associated with their role in Cl⁻ secretion. The cAMP- and Ca²⁺-dependent secretory responses were inhibited by the removal of HCO₃ ⁻ suggesting that these responses were due to electrogenic HCO₃ ⁻ secretion. We conclude that the ileum of the possum does not secrete Cl⁻ due to low levels of NKCC1 expression. It does however appear to secrete HCO₃ ⁻. These results are further significant examples of differences in the transport function of the possum intestinal epithelium compared with eutherian mammals.     

Phosphorylation-Mediated Control of Histone Chaperone ASF1 Levels by Tousled-Like Kinases

Histone chaperones are at the hub of a diverse interaction networks integrating a plethora of chromatin modifying activities. Histone H3/H4 chaperone ASF1 is a target for cell-cycle regulated Tousled-like kinases (TLKs) and both proteins cooperate during chromatin replication. However, the precise role of post-translational modification of ASF1 remained unclear. Here, we identify the TLK phosphorylation sites for both Drosophila and human ASF1 proteins. Loss of TLK-mediated phosphorylation triggers hASF1a and dASF1 degradation by proteasome-dependent and independent mechanisms respectively. Consistent with this notion, introduction of phosphorylation-mimicking mutants inhibits hASF1a and dASF1 degradation. Human hASF1b is also targeted for proteasome-dependent degradation, but its stability is not affected by phosphorylation indicating that other mechanisms are likely to be involved in control of hASF1b levels. Together, these results suggest that ASF1 cellular levels are tightly controlled by distinct pathways and provide a molecular mechanism for post-translational regulation of dASF1 and hASF1a by TLK kinases.     

Natural Selection of Human Embryos: Decidualizing Endometrial Stromal Cells Serve as Sensors of Embryo Quality upon Implantation

Background

Pregnancy is widely viewed as dependent upon an intimate dialogue, mediated by locally secreted factors between a developmentally competent embryo and a receptive endometrium. Reproductive success in humans is however limited, largely because of the high prevalence of chromosomally abnormal preimplantation embryos. Moreover, the transient period of endometrial receptivity in humans uniquely coincides with differentiation of endometrial stromal cells (ESCs) into highly specialized decidual cells, which in the absence of pregnancy invariably triggers menstruation. The role of cyclic decidualization of the endometrium in the implantation process and the nature of the decidual cytokines and growth factors that mediate the crosstalk with the embryo are unknown.

Methodology/Principal Findings

We employed a human co-culture model, consisting of decidualizing ESCs and single hatched blastocysts, to identify the soluble factors involved in implantation. Over the 3-day co-culture period, approximately 75% of embryos arrested whereas the remainder showed normal development. The levels of 14 implantation factors secreted by the stromal cells were determined by multiplex immunoassay. Surprisingly, the presence of a developing embryo had no significant effect on decidual secretions, apart from a modest reduction in IL-5 levels. In contrast, arresting embryos triggered a strong response, characterized by selective inhibition of IL-1β, -6, -10, -17, -18, eotaxin, and HB-EGF secretion. Co-cultures were repeated with undifferentiated ESCs but none of the secreted cytokines were affected by the presence of a developing or arresting embryo.

Conclusions

Human ESCs become biosensors of embryo quality upon differentiation into decidual cells. In view of the high incidence of gross chromosomal errors in human preimplantation embryos, cyclic decidualization followed by menstrual shedding may represent a mechanism of natural embryo selection that limits maternal investment in developmentally impaired pregnancies.     

Natural Selection of Human Embryos: Impaired Decidualization of Endometrium Disables Embryo-Maternal Interactions and Causes Recurrent Pregnancy Loss

Background

Recurrent pregnancy loss (RPL), defined as 3 or more consecutive miscarriages, is widely attributed either to repeated chromosomal instability in the conceptus or to uterine factors that are poorly defined. We tested the hypothesis that abnormal cyclic differentiation of endometrial stromal cells (ESCs) into specialized decidual cells predisposes to RPL, based on the observation that this process may not only be indispensable for placenta formation in pregnancy but also for embryo recognition and selection at time of implantation.

Methodology/Principal Findings

Analysis of mid-secretory endometrial biopsies demonstrated that RPL is associated with decreased expression of the decidual marker prolactin (PRL) but increased levels of prokineticin-1 (PROK1), a cytokine that promotes implantation. These in vivo findings were entirely recapitulated when ESCs were purified from patients with and without a history of RPL and decidualized in culture. In addition to attenuated PRL production and prolonged and enhanced PROK1 expression, RPL was further associated with a complete dysregulation of both markers upon treatment of ESC cultures with human chorionic gonadotropin, a glycoprotein hormone abundantly expressed by the implanting embryo. We postulated that impaired embryo recognition and selection would clinically be associated with increased fecundity, defined by short time-to-pregnancy (TTP) intervals. Woman-based analysis of the mean and mode TTP in a cohort of 560 RPL patients showed that 40% can be considered “superfertile”, defined by a mean TTP of 3 months or less.

Conclusions

Impaired cyclic decidualization of the endometrium facilitates implantation yet predisposes to subsequent pregnancy failure by disabling natural embryo selection and by disrupting the maternal responses to embryonic signals. These findings suggest a novel pathological pathway that unifies maternal and embryonic causes of RPL.     

Further analysis of the involvement of the envelope anion channel PIRAC in chloroplast protein import.

The ability of preferredoxin to inactivate a 50-pS anion channel of the chloroplast inner membrane in the presence of an energy source was investigated using single-channel recordings. It was found that preferredoxin cannot inactivate the channel when GTP is the only energy source present. From this it is concluded that the precursor has to interact with the, translocon of the inner membrane of chloroplasts (Tic) complex to be able to inactivate the 50-pS anion channel. The ability of two mutants of preferredoxin with deletions in their transit sequence to inactivate the channel was also tested. Both mutants have been shown to have a similar binding affinity for the chloroplast envelope, but only one is able to fully translocate. The mutants were both able to inactivate the channel in a similar manner. From this it is concluded that full translocation is not necessary for the inactivation of the channel. It is also shown that preferredoxin is capable of inactivating the 50-pS anion channel in the chloroplast-attached configuration as was previously found in the inside-out configuration. From this it is concluded that stromal factors do not influence the protein-import-induced inactivation of the 50-pS anion channel of the chloroplast inner membrane. Finally the effect of the anion channel blocker 4, 4'-diisothiocyanostilbene-2,2'-disulfonate (DIDS) on the channel activity and on protein import was investigated. It was found that DIDS blocked the channel. Furthermore the addition of the channel blocker reduces the efficiency of import to 52%. This leads to the conclusion that correct functioning of the channel is important for protein import.     

Interdecadal trends in composition,density, size,and mean trophic level of fish species and guilds before and after coastal development in the Mexican Caribbean

This study explores the possible influence of human coastal development (before and after) and protected area status (within and outside a marine protected area, MPA) on composition, density, and maximum size of fish species and guilds, including mean trophic level of the fish community, in four localities of the Yucatan Peninsula, Mexican Caribbean. Reef fish density, maximum length, species composition, and trophic guilds were recorded by SCUBA belt transects and stationary points in fore reef and lagoon reef areas at decadal intervals (1995–1998, 2006–2010, 2014–2015). Mean density of most species and guilds decreased significantly through the years, as also did mean trophic level of the fish community. Some fish species increased in length. Fish density for many species was larger outside than inside the MPA in 1995–1998; however, the difference tended to disappear in the more recent decades, which reflects either a positive effect of the MPA, or a detrimental effect of coastal development in the non-protected area. Nevertheless, the overall negative trends suggest a regional or global rather than a local cause.     

The impact of chronic environmental stressors on growing pigs, Sus scrofa (Part 2): social behaviour

The effects of common and concurrent environmental stressors on the social behaviour of farm animals are poorly understood. Here, we report the results of a multifactorial experiment designed specifically to examine the individual, additive or interactive effects of elevated ammonia, noise and low light (LL) levels on the social behaviour of growing pigs. Social behaviour was measured in terms of the nature, frequency and duration of both initiated and response behaviours for 4 weeks following mixing of the groups. General activity patterns, group cohesion and social discrimination were also examined as a function of the environmental treatments. Elevated concentrations of atmospheric ammonia (～20 v. <5 ppm) and LL intensity (～40 v. 200 lux) had the most pronounced effects, particularly on the nature of social interactions, with pigs under these conditions showing more aggression in the early stages of the experiment. In addition, pigs exposed to a high level of mechanical noise representative of artificial ventilation (～80 v. 40 dB [A]) were less submissive to aggressive acts, while pigs in ～20 ppm ammonia showed more reciprocated aggression when in coincident LL (<40 lux). The results indicate that atmospheric ammonia at commonly experienced concentrations may undermine social stability, particularly in the presence of low lighting, though the mechanisms are currently unknown. These findings have implications for the welfare of growing pigs and hence policy makers and farmers alike, with respect to the improvement of welfare in intensive pig farming.