Siah2 control of T-regulatory cells limits anti-tumor immunity.

Understanding the mechanisms underlying anti-tumor immunity is pivotal for improving immune-based cancer therapies. Here, we report that growth of BRAF-mutant melanoma cells is inhibited, up to complete rejection, in Siah2-/- mice. Growth-inhibited tumors exhibit increased numbers of intra-tumoral activated T cells and decreased expression of Ccl17, Ccl22, and Foxp3. Marked reduction in Treg proliferation and tumor infiltration coincide with G1 arrest in tumor infiltrated Siah2-/- Tregs in vivo or following T cell stimulation in culture, attributed to elevated expression of the cyclin-dependent kinase inhibitor p27, a Siah2 substrate. Growth of anti-PD-1 therapy resistant melanoma is effectively inhibited in Siah2-/- mice subjected to PD-1 blockade, indicating synergy between PD-1 blockade and Siah2 loss. Low SIAH2 and FOXP3 expression is identified in immune responsive human melanoma tumors. Overall, Siah2 regulation of Treg recruitment and cell cycle progression effectively controls melanoma development and Siah2 loss in the host sensitizes melanoma to anti-PD-1 therapy

Notch signaling regulates metabolic heterogeneity in glioblastoma stem cells.

Glioblastoma (GBM) stem cells (GSCs) reside in both hypoxic and vascular microenvironments within tumors. The molecular mechanisms that allow GSCs to occupy such contrasting niches are not understood. We used patient-derived GBM cultures to identify GSC subtypes with differential activation of Notch signaling, which co-exist in tumors but occupy distinct niches and match their metabolism accordingly. Multipotent GSCs with Notch pathway activation reside in perivascular niches, and are unable to entrain anaerobic glycolysis during hypoxia. In contrast, most CD133-expressing GSCs do not depend on canonical Notch signaling, populate tumors regardless of local vascularity and selectively utilize anaerobic glycolysis to expand in hypoxia. Ectopic activation of Notch signaling in CD133-expressing GSCs is sufficient to suppress anaerobic glycolysis and resistance to hypoxia. These findings demonstrate a novel role for Notch signaling in regulating GSC metabolism and suggest intratumoral GSC heterogeneity ensures metabolic adaptations to support tumor growth in diverse tumor microenvironments

Regulation of transcriptional elongation in pluripotency and cell differentiation by the PHD-finger protein Phf5a

Pluripotent embryonic stem cells (ESCs) self-renew or differentiate into all tissues of the developing embryo and cell-specification factors are necessary to balance gene expression. Here we delineate the function of the PHD-finger protein 5a (Phf5a) in ESC self-renewal and ascribe its role in regulating pluripotency, cellular reprogramming, and myoblast specification. We demonstrate that Phf5a is essential for maintaining pluripotency, since depleted ESCs exhibit hallmarks of differentiation. Mechanistically, we attribute Phf5a function to the stabilization of the Paf1 transcriptional complex and control of RNA polymerase II elongation on pluripotency loci. Apart from an ESC-specific factor, we demonstrate that Phf5a controls differentiation of adult myoblasts. Our findings suggest a potent mode of regulation by the Phf5a in stem cells, which directs their transcriptional program ultimately regulating maintenance of pluripotency and cellular reprogramming

Calorie Restriction Suppresses Age-Dependent Hippocampal Transcriptional Signatures.

Calorie restriction (CR) enhances longevity and mitigates aging phenotypes in numerous species. Physiological responses to CR are cell-type specific and variable throughout the lifespan. However, the mosaic of molecular changes responsible for CR benefits remains unclear, particularly in brain regions susceptible to deterioration during aging. We examined the influence of long-term CR on the CA1 hippocampal region, a key learning and memory brain area that is vulnerable to age-related pathologies, such as Alzheimer's disease (AD). Through mRNA sequencing and NanoString nCounter analysis, we demonstrate that one year of CR feeding suppresses age-dependent signatures of 882 genes functionally associated with synaptic transmission-related pathways, including calcium signaling, long-term potentiation (LTP), and Creb signaling in wild-type mice. By comparing the influence of CR on hippocampal CA1 region transcriptional profiles at younger-adult (5 months, 2.5 months of feeding) and older-adult (15 months, 12.5 months of feeding) timepoints, we identify conserved upregulation of proteome quality control and calcium buffering genes, including heat shock 70 kDa protein 1b (Hspa1b) and heat shock 70 kDa protein 5 (Hspa5), protein disulfide isomerase family A member 4 (Pdia4) and protein disulfide isomerase family A member 6 (Pdia6), and calreticulin (Calr). Expression levels of putative neuroprotective factors, klotho (Kl) and transthyretin (Ttr), are also elevated by CR in adulthood, although the global CR-specific expression profiles at younger and older timepoints are highly divergent. At a previously unachieved resolution, our results demonstrate conserved activation of neuroprotective gene signatures and broad CR-suppression of age-dependent hippocampal CA1 region expression changes, indicating that CR functionally maintains a more youthful transcriptional state within the hippocampal CA1 sector

Biological variance within mRNA sequencing condition groups.

<p>The squared coefficient of variation (CV²) was plotted against Fragments Per Kilobase of exon per Million fragments mapped (log₁₀FPKM), representing the total distribution of mRNA sequence reads for each condition group, which are depicted as follows, 5 months AL (coral), 5 months CR (green), 15 months AL (blue), and 15 months CR (purple).</p

Differential gene expression via mRNA sequence analysis pairwise comparisons.

<p>The total number of differentially expressed transcripts identified by total mRNA sequencing within each pairwise comparison are indicated, along with the log₂ fold change (Log2FC) range and the number of upregulated (gray) and downregulated (white) targets (p<0.01, q<0.05).</p

CR upregulation of neuroprotective gene signatures is conserved at 5 and 15 months, despite divergent transcriptional profiles.

<p>(A) 102 genes are common to both the 15 and 5 month CR vs. AL differential expression mRNA sequencing profiles. Of the 102 common genes, 70/102 (69%) of significant changes occur in the opposite direction. (B) Normalized age- and diet-dependent FPKM levels for select protein folding and calcium buffering genes (*p<0.01, q<0.05); heat shock 70 kDa protein 5 (Hspa5), heat shock 70 kDa protein 1B (Hspa1b), calreticulin (Calr). (C) Log₂ fold change levels for the 32 conserved gene expression changes within 15 (blue) and 5 (red) month CR vs. AL expression profiles that occurred in the same direction are depicted (p<0.01, q<0.05).</p

Average body weight following sustained 30% CR or AL feeding.

<p>Beginning at approximately 2.5 months of age, wild-type female mice were maintained on 30% CR (purple) or AL (blue) diets and were sacrificed following 2.5 or 12.5 months of diet administration. Body weight was measured approximately twice weekly. Within the first 2 weeks of AL and CR feeding, mice maintained on the 30% CR diet lost an average of 12% of their body weight (t-test, p<0.001), which stabilized for the remainder of the study. For study days 1–85, n = 12–18, and for study days 85–365, n = 6–14, reflecting sacrifice of the first group of mice at 5 months of age; mean +/- SEM.</p

H3K27me3 dynamics dictate evolving uterine states in pregnancy and parturition.

Uncovering the causes of pregnancy complications such as preterm labor requires greater insight into how the uterus remains in a noncontractile state until term and then surmounts this state to enter labor. Here, we show that dynamic generation and erasure of the repressive histone modification tri-methyl histone H3 lysine 27 (H3K27me3) in decidual stromal cells dictate both elements of pregnancy success in mice. In early gestation, H3K27me3-induced transcriptional silencing of select gene targets ensured uterine quiescence by preventing the decidua from expressing parturition-inducing hormone receptors, manifesting type 1 immunity, and most unexpectedly, generating myofibroblasts and associated wound-healing responses. In late gestation, genome-wide H3K27 demethylation allowed for target gene upregulation, decidual activation, and labor entry. Pharmacological inhibition of H3K27 demethylation in late gestation not only prevented term parturition, but also inhibited delivery while maintaining pup viability in a noninflammatory model of preterm parturition. Immunofluorescence analysis of human specimens suggested that similar regulatory events might occur in the human decidua. Together, these results reveal the centrality of regulated gene silencing in the uterine adaptation to pregnancy and suggest new areas in the study and treatment of pregnancy disorders