PI3K/mTOR signaling regulates prostatic branching morphogenesis

Prostatic branching morphogenesis is an intricate event requiring precise temporal and spatial integration of numerous hormonal and growth factor-regulated inputs, yet relatively little is known about the downstream signaling pathways that orchestrate this process. In this study, we use a novel mesenchyme-free embryonic prostate culture system, newly available mTOR inhibitors and a conditional PTEN loss-of-function model to investigate the role of the interconnected PI3K and mTOR signaling pathways in prostatic organogenesis. We demonstrate that PI3K levels and PI3K/mTOR activity are robustly induced by androgen during murine prostatic development and that PI3K/mTOR signaling is necessary for prostatic epithelial bud invasion of surrounding mesenchyme. To elucidate the cellular mechanism by which PI3K/mTOR signaling regulates prostatic branching, we show that PI3K/mTOR inhibition does not significantly alter epithelial proliferation or apoptosis, but rather decreases the efficiency and speed with which the developing prostatic epithelial cells migrate. Using mTOR kinase inhibitors to tease out the independent effects of mTOR signaling downstream of PI3K, we find that simultaneous inhibition of mTORC1 and mTORC2 activity attenuates prostatic branching and is sufficient to phenocopy combined PI3K/mTOR inhibition. Surprisingly, however, mTORC1 inhibition alone has the reverse effect, increasing the number and length of prostatic branches. Finally, simultaneous activation of PI3K and downstream mTORC1/C2 via epithelial PTEN loss-of-function also results in decreased budding reversible by mTORC1 inhibition, suggesting that the effect of mTORC1 on branching is not primarily mediated by negative feedback on PI3K/mTORC2 signaling. Taken together, our data point to an important role for PI3K/mTOR signaling in prostatic epithelial invasion and migration and implicates the balance of PI3K and downstream mTORC1/C2 activity as a critical regulator of prostatic epithelial morphogenesis.     

A hormone-dependent module regulating energy balance

Under fasting conditions, metazoans maintain energy balance by shifting from glucose to fat burning. In the fasted state, SIRT1 promotes catabolic gene expression by deacetylating the forkhead factor FOXO in response to stress and nutrient deprivation. The mechanisms by which hormonal signals regulate FOXO deacetylation remain unclear, however. We identified a hormone-dependent module, consisting of the Ser/Thr kinase SIK3 and the class IIa deacetylase HDAC4, which regulates FOXO activity in Drosophila. During feeding, HDAC4 is phosphorylated and sequestered in the cytoplasm by SIK3, whose activity is upregulated in response to insulin. SIK3 is inactivated during fasting, leading to the dephosphorylation and nuclear translocation of HDAC4 and to FOXO deacetylation. SIK3 mutant flies are starvation sensitive, reflecting FOXO-dependent increases in lipolysis that deplete triglyceride stores; reducing HDAC4 expression restored lipid accumulation. Our results reveal a hormone-regulated pathway that functions in parallel with the nutrient-sensing SIRT1 pathway to maintain energy balance.     

Raloxifene attenuates Gas6 and apoptosis in experimental aortic valve disease in renal failure

Renal failure is associated with aortic valve calcification. Using our rat model of uremia-induced reversible aortic valve calcification, we assessed the role of apoptosis and survival pathways in that disease. We also explored the effects of raloxifene, an estrogen receptor modulator, on valvular calcification. Gene array analysis was performed in aortic valves obtained from three groups of rats (n = 7 rats/group): calcified valves obtained from rats fed with uremic diet, valves after calcification resolution following diet cessation, and control. In addition, four groups of rats (n = 10 rats/group) were used to evaluate the effect of raloxifene in aortic valve calcification: three groups as mentioned above and a fourth group fed with the uremic diet that also received daily raloxifene. Evaluation included imaging, histology, and antigen expression analysis. Gene array results showed that the majority of the altered expressed genes were in diet group valves. Most apoptosis-related genes were changed in a proapoptotic direction in calcified valves. Apoptosis and decreases in several survival pathways were confirmed in calcified valves. Resolution of aortic valve calcification was accompanied by decreased apoptosis and upregulation of survival pathways. Imaging and histology demonstrated that raloxifene significantly decreased aortic valve calcification. In conclusion, downregulation of several survival pathways and apoptosis are involved in the pathogenesis of aortic valve calcification. The beneficial effect of raloxifene in valve calcification is related to apoptosis modulation. This novel observation is important for developing remedies for aortic valve calcification in patients with renal failure.     

Female ixodid ticks grow endocuticle during the rapid phase of engorgement

Lees (Proc Zool Soc Lond 121:759–772, 1952) concluded that the ixodid tick Ixodes ricinus grows endocuticle during the slow but not during the rapid, phase of engorgement, a conclusion supported by Andersen and Roepstorff (Insect Biochem Mol Biol 35:1181–1188, 2005) for the same species. In this study analysis of dimensional data and cuticle weight measurements from female ixodid ticks (Amblyomma hebraeum) were used to test this hypothesis. Both approaches showed that endocuticle growth continues during the rapid phase, tapering to zero at a fed/unfed weight ratio of ~60. Of the total mass of cuticle in the engorged tick 32–43% was formed during the rapid phase. We demonstrate that if cuticle growth stopped at the end of the slow phase, there would not be sufficient cuticle to account for the thickness of cuticle observed at the end of engorgement. This finding is consistent with prior studies of Rhipicephalus (Boophilus) microplus, and with a dimensional analysis of the cuticle thickness data of Lees for I. ricinus, in contradiction to his conclusion from an analysis of tick cuticle weight measurements. An examination of cuticle weight measurements for I. ricinus by Andersen and Roepstorff similarly supports the finding of cuticle growth during the rapid phase. All ixodid ticks undergo major body expansion, typically tenfold or more, during a rapid phase of engorgement and require sufficient cuticle at the end of that process to contain their body. The fact that cuticle grows during the rapid phase of engorgement in three species suggests that this is a general characteristic of the family Ixodidae.     

Type III IFNs in pteropid bats: differential expression patterns provide evidence for distinct roles in antiviral immunity

Bats are known to harbor a number of emerging and re-emerging zoonotic viruses, many of which are highly pathogenic in other mammals but result in no clinical symptoms in bats. The ability of bats to coexist with viruses may be the result of rapid control of viral replication early in the immune response. IFNs provide the first line of defense against viral infection in vertebrates. Type III IFNs (IFN-λs) are a recently identified IFN family that share similar antiviral activities with type I IFNs. To our knowledge, we demonstrate the first functional analysis of type III IFNs from any species of bat, with the investigation of two IFN-λ genes from the pteropid bat, Pteropus alecto. Our results demonstrate that bat type III IFN has similar antiviral activity to type I and III IFNs from other mammals. In addition, the two bat type III IFNs are differentially induced relative to each other and to type I IFNs after treatment or transfection with synthetic dsRNA. Infection with the bat paramyxovirus, Tioman virus, resulted in no upregulation of type I IFN production in bat splenocytes but was capable of inducing a type III IFN response in three of the four bats tested. To our knowledge, this is the first report to describe the simultaneous suppression of type I IFN and induction of type III IFN after virus infection. These results may have important implications for the role of type III IFNs in the ability of bats to coexist with viruses.     

Human and rhesus APOBEC3D, APOBEC3F, APOBEC3G, and APOBEC3H demonstrate a conserved capacity to restrict Vif-deficient HIV-1

Successful intracellular pathogens must evade or neutralize the innate immune defenses of their host cells and render the cellular environment permissive for replication. For example, to replicate efficiently in CD4(+) T lymphocytes, human immunodeficiency virus type 1 (HIV-1) encodes a protein called viral infectivity factor (Vif) that promotes pathogenesis by triggering the degradation of the retrovirus restriction factor APOBEC3G. Other APOBEC3 proteins have been implicated in HIV-1 restriction, but the relevant repertoire remains ambiguous. Here we present the first comprehensive analysis of the complete, seven-member human and rhesus APOBEC3 families in HIV-1 restriction. In addition to APOBEC3G, we find that three other human APOBEC3 proteins, APOBEC3D, APOBEC3F, and APOBEC3H, are all potent HIV-1 restriction factors. These four proteins are expressed in CD4(+) T lymphocytes, are packaged into and restrict Vif-deficient HIV-1 when stably expressed in T cells, mutate proviral DNA, and are counteracted by HIV-1 Vif. Furthermore, APOBEC3D, APOBEC3F, APOBEC3G, and APOBEC3H of the rhesus macaque also are packaged into and restrict Vif-deficient HIV-1 when stably expressed in T cells, and they are all neutralized by the simian immunodeficiency virus Vif protein. On the other hand, neither human nor rhesus APOBEC3A, APOBEC3B, nor APOBEC3C had a significant impact on HIV-1 replication. These data strongly implicate a combination of four APOBEC3 proteins--APOBEC3D, APOBEC3F, APOBEC3G, and APOBEC3H--in HIV-1 restriction.     

Phosphorylation directly regulates the intrinsic DNA cytidine deaminase activity of activation-induced deaminase and APOBEC3G protein

The beneficial effects of DNA cytidine deamination by activation-induced deaminase (AID; antibody gene diversification) and APOBEC3G (retrovirus restriction) are tempered by probable contributions to carcinogenesis. Multiple regulatory mechanisms serve to minimize this detrimental outcome. Here, we show that phosphorylation of a conserved threonine attenuates the intrinsic activity of activation-induced deaminase (Thr-27) and APOBEC3G (Thr-218). Phospho-null alanine mutants maintain intrinsic DNA deaminase activity, whereas phospho-mimetic glutamate mutants are inactive. The phospho-mimetic variants fail to mediate isotype switching in activated mouse splenic B lymphocytes or suppress HIV-1 replication in human T cells. Our data combine to suggest a model in which this critical threonine acts as a phospho-switch that fine-tunes the adaptive and innate immune responses and helps protect mammalian genomic DNA from procarcinogenic lesions.     

Developmental dynamics of myogenesis in the shipworm <Emphasis Type="Italic">Lyrodus pedicellatus</Emphasis> (Mollusca: Bivalvia)

Background

The shipworm Lyrodus pedicellatus is a wood-boring bivalve with an unusual vermiform body. Although its larvae are brooded, they retain the general appearance of a typical bivalve veliger-type larva. Here, we describe myogenesis of L. pedicellatus revealed by filamentous actin labelling and discuss the data in a comparative framework in order to test for homologous structures that might be part of the bivalve (larval) muscular ground pattern.

Results

Five major muscle systems were identified: a velum retractor, foot retractor, larval retractor, a distinct mantle musculature and an adductor system. For a short period of larval life, an additional ventral larval retractor is present. Early in development, a velum muscle ring and an oral velum musculature emerge. In late stages the lateral and dorsal mantle musculature, paired finger-shaped muscles, an accessory adductor and a pedal plexus are formed. Similar to other bivalve larvae, L. pedicellatus exhibits three velum retractor muscles, but in contrast to other species, one of them disappears in early stages of L. pedicellatus. The remaining two velum retractors are considerably remodelled during late larval development and are most likely incorporated into the elaborate mantle musculature of the adult.

Conclusions

To our knowledge, this is the first account of any larval retractor system that might contribute to the adult bodyplan of a (conchiferan) mollusk. A comparative analysis shows that a pedal plexus, adductors, a larval velum ring, velum retractors and a ventral larval retractor are commonly found among bivalve larvae, and thus most likely belong to the ground pattern of the bivalve larval musculature.