The p57 CDKi integrates stress signals into cell-cycle progression to promote cell survival upon stress.

The p57(Kip2) cyclin-dependent kinase inhibitor (CDKi) has been implicated in embryogenesis, stem-cell senescence and pathologies, but little is known of its role in cell cycle control. Here, we show that p57(Kip2) is targeted by the p38 stress-activated protein kinase (SAPK). Phosphorylation of p57(Kip2) at T143 by p38 enhances its association with and inhibition of Cdk2, which results in cell-cycle delay upon stress. Genetic inactivation of the SAPK or the CDKi abolishes cell-cycle delay upon osmostress and results in decreased cell viability. Oxidative stress and ionomycin also induce p38-mediated phosphorylation of p57 and cells lacking p38 or p57 display reduced viability to these stresses. Therefore, cell survival to various stresses depends on p57 phosphorylation by p38 that inhibits CDK activity. Together, these findings provide a novel molecular mechanism by which cells can delay cell cycle progression to maximize cell survival upon stress.

Trabecular bone score in the hip: a new method to examine hip bone microarchitecture-a feasibility study.

Our study found, in older adults who are residents of long-term care facilities, assessing hip microarchitecture with DXA-derived bone texture score may serve as a supplement to bone mineral density to improve fracture prediction and to facilitate decision-making for pharmacological management. PURPOSE: Many patients with high fragility fracture risk do not have a sufficiently low bone mineral density (BMD) to become eligible for osteoporosis treatment. They often have deteriorated bone microarchitecture despite a normal or only mildly abnormal BMD. We sought to examine the beta version of the trabecular bone score (TBS) algorithm for the hip: TBS Hip, an indirect index of bone microarchitecture, and assess if TBS Hip brings complementary information to other bone quality indices such as BMD and bone turnover markers (BTMs) to further improve identifying individuals who are at high risk for fractures. METHODS: In this analysis, we considered baseline TBS Hip at total hip, femoral neck, and greater trochanter, TBS at lumbar spine, BMD at all of these skeletal sites, and BTMs in 132 postmenopausal women who were residents of long-term care (LTC) facilities enrolled in a randomized placebo-controlled osteoporosis clinical trial. RESULTS: On average, participants were 85.2 years old and had a BMI of 26.9 kg/m2. The correlation coefficient between BMD and TBS Hip at total hip, femoral neck, and greater trochanter was 0.50, 0.32, and 0.39 respectively (all p < 0.0001). The correlation coefficient between BMD and lumbar spine TBS was 0.52 (p < 0.0001). There was no statistically significant correlation between BTMs with TBS at lumbar spine or TBS Hip at total hip, femoral neck, and greater trochanter. CONCLUSION: Among older women residing in LTC facilities, there was a moderate correlation between measures of BMD and TBS Hip at total hip, femoral neck, and greater trochanter, suggesting TBS Hip may provide complementary information to BMD .

The p38 SAPK is recruited to chromatin via its interaction with transcription factors.

In mammals, the stress-activated protein kinase (SAPK) p38 coordinates a rapid and complex transcriptional program to adapt to sudden changes in the extracellular environment. Although a number of genes have been reported to be under the control of p38, the basic mechanisms of transcriptional regulation by this SAPK remain uncharacterized. Here we show that in response to osmotic shock, anisomycin- or TNFα-activated p38 SAPK is recruited to stress-induced genes. The MAPKK MKK6 is also found at stress-responsive promoters. The recruitment of RNA polymerase II complex to the target promoters requires p38 activity. Moreover, when tethered to DNA as a LexA fusion protein, p38 activates transcription in a stress-regulated manner. Thus, p38 activity allows for recruitment of RNA polymerase and transcription initiation. p38 directly phosphorylates and interacts with the transcription factor Elk1. p38 activity is necessary for the recruitment of Elk1 to the c-Fos promoter, and knocking down Elk1 by siRNAs compromises both p38 recruitment to the c-Fos promoter and c-Fos transcriptional up-regulation upon osmostress. In addition, p38 recruitment to the osmoinducible gene Cox2 and the TNFα target gene IL8 is mediated by the transcription factors AP1 and NFκB, respectively. Therefore, anchoring of active SAPK to target genes is mediated by transcription factors. The presence of active p38 at open reading frames also suggests the involvement of the SAPK in elongation. Taken together, SAPK recruitment to target genes appears to be a broad mechanism to regulate transcription that has been preserved from yeast to mammals.

In Vivo Follow-Up of Gene Inhibition in Solid Tumors Using Peptide-Based Nanoparticles for siRNA Delivery.

Small interfering RNA (siRNA) exhibits a high degree of specificity for targeting selected genes. They are efficient on cells in vitro, but in vivo siRNA therapy remains a challenge for solid tumor treatment as siRNAs display difficulty reaching their intracellular target. The present study was designed to show the in vivo efficiency of a new peptide (WRAP5), able to form peptide-based nanoparticles (PBN) that can deliver siRNA to cancer cells in solid tumors. WRAP5:siRNA nanoparticles targeting firefly luciferase (Fluc) were formulated and assayed on Fluc-expressing U87 glioblastoma cells. The mode of action of WRAP5:siRNA by RNA interference was first confirmed in vitro and then investigated in vivo using a combination of bioluminescent reporter genes. Finally, histological analyses were performed to elucidate the cell specificity of this PBN in the context of brain tumors. In vitro and in vivo results showed efficient knock-down of Fluc expression with no toxicity. WRAP5:siFluc remained in the tumor for at least 10 days in vivo. Messenger RNA (mRNA) analyses indicated a specific decrease in Fluc mRNA without affecting tumor growth. Histological studies identified PBN accumulation in the cytoplasm of tumor cells but also in glial and neuronal cells. Through in vivo molecular imaging, our findings established the proof of concept for specific gene silencing in solid tumors. The evidence generated could be translated into therapy for any specific gene in different types of tumors without cell type specificity but with high molecular specificity.

Whole genome analysis of p38 SAPK-mediated gene expression upon stress.

BACKGROUND: Cells have the ability to respond and adapt to environmental changes through activation of stress-activated protein kinases (SAPKs). Although p38 SAPK signalling is known to participate in the regulation of gene expression little is known on the molecular mechanisms used by this SAPK to regulate stress-responsive genes and the overall set of genes regulated by p38 in response to different stimuli. RESULTS: Here, we report a whole genome expression analyses on mouse embryonic fibroblasts (MEFs) treated with three different p38 SAPK activating-stimuli, namely osmostress, the cytokine TNFalpha and the protein synthesis inhibitor anisomycin. We have found that the activation kinetics of p38alpha SAPK in response to these insults is different and also leads to a complex gene pattern response specific for a given stress with a restricted set of overlapping genes. In addition, we have analysed the contribution of p38alpha the major p38 family member present in MEFs, to the overall stress-induced transcriptional response by using both a chemical inhibitor (SB203580) and p38alpha deficient (p38alpha-/-) MEFs. We show here that p38 SAPK dependency ranged between 60% and 88% depending on the treatments and that there is a very good overlap between the inhibitor treatment and the ko cells. Furthermore, we have found that the dependency of SAPK varies depending on the time the cells are subjected to osmostress. CONCLUSIONS: Our genome-wide transcriptional analyses shows a selective response to specific stimuli and a restricted common response of up to 20% of the stress up-regulated early genes that involves an important set of transcription factors, which might be critical for either cell adaptation or preparation for continuous extra-cellular changes. Interestingly, up to 85% of the up-regulated genes are under the transcriptional control of p38 SAPK. Thus, activation of p38 SAPK is critical to elicit the early gene expression program required for cell adaptation to stress.

Skeletal Anomalies in Senegalese Sole (Solea senegalensis, Kaup) Fed with Different Commercial Enriched Artemia: A Study in Postlarvae and Juveniles.

The high incidence of skeletal anomalies in Senegalese sole (Solea senegalensis) still constitutes a bottleneck constraining its production. There are diverse commercially available products for the enrichment of live preys, but few reports of their influence on skeletogenesis in Senegalese sole. This study evaluated the presence of vertebral anomalies in postlarvae and juvenile Senegalese sole fed with Artemia spp. metanauplii enriched with four commercial products (EA, EB, EC, and ED) in a fish farm. The most frequent alterations consisted of deformations of the neural/haemal arches and spines and fusions and deformations of hypurals, epural, or parhypural. The correspondence analysis ordered fish from each age in separated semiaxis, indicating the presence of different anomaly patterns for the two sampled stages. The results showed only very light changes in the frequency of vertebral abnormalities among tested enrichment products, i.e., individuals from EC and EA lots displayed less vertebral body anomalies and/or vertebral column deviations at 31 and 105 days after hatching, respectively. The existence of a large shared malformation pattern in all the experimental groups leads to impute to the rearing conditions as the main driving factor of the onset of such group of anomalies, probably masking some dietary effect.

Identification of the major sex-determining region of turbot (Scophthalmus maximus).

Sex determination in fish is a labile character in evolutionary terms. The sex-determining (SD) master gene can differ even between closely related fish species. This group is an interesting model for studying the evolution of the SD region and the gonadal differentiation pathway. The turbot (Scophthalmus maximus) is a flatfish of great commercial value, where a strong sexual dimorphism exists for growth rate. Following a QTL and marker association approach in five families and a natural population, we identified the main SD region of turbot at the proximal end of linkage group (LG) 5, close to the SmaUSC-E30 marker. The refined map of this region suggested that this marker would be 2.6 cM and 1.4 Mb from the putative SD gene. This region appeared mostly undifferentiated between males and females, and no relevant recombination frequency differences were detected between sexes. Comparative genomics of LG5 marker sequences against five model species showed no similarity of this chromosome to the sex chromosomes of medaka, stickleback, and fugu, but suggested a similarity to a sex-associated QTL from Oreochromis spp. The segregation analysis of the closest markers to the SD region demonstrated a ZW/ZZ model of sex determination in turbot. A small proportion of families did not fit perfectly with this model, which suggests that other minor genetic and/or environmental factors are involved in sex determination in this species.