Elevated CO2 levels (hypercapnia) frequently occur in patients with obstructive pulmonary diseases and are associated with increased mortality. However, the effects of hypercapnia on non-neuronal tissues and the mechanisms that mediate these effects are largely unknown. Here, we develop Drosophila as a genetically tractable model for defining non-neuronal CO 2 responses and response pathways. We show that hypercapnia significantly impairs embryonic morphogenesis, egg laying, and egg hatching even in mutants lacking the Gr63a neuronal CO 2 sensor. Consistent with previous reports that hypercapnic acidosis can suppress mammalian NF-B-regulated innate immune genes, we find that in adult flies and the phagocytic immune-responsive S2* cell line, hypercapnia suppresses induction of specific antimicrobial peptides that are regulated by Relish, a conserved Rel/NF-B family member. Correspondingly, modest hypercapnia (7-13%) increases mortality of flies inoculated with E. faecalis, A. tumefaciens, or S. aureus. During E. faecalis and A. tumefaciens infection, increased bacterial loads were observed, indicating that hypercapnia can decrease host resistance. Hypercapnic immune suppression is not mediated by acidosis, the olfactory CO 2 receptor Gr63a, or by nitric oxide signaling. Further, hypercapnia does not induce responses characteristic of hypoxia, oxidative stress, or heat shock. Finally, proteolysis of the Relish IB-like domain is unaffected by hypercapnia, indicating that immunosuppression acts downstream of, or in parallel to, Relish proteolytic activation. Our results suggest that hypercapnic immune suppression is mediated by a conserved response pathway, and illustrate a mechanism by which hypercapnia could contribute to worse outcomes of patients with advanced lung disease, who frequently suffer from both hypercapnia and respiratory infections.COPD ͉ hypercapnia ͉ Relish ͉ NF-B ͉ Gr63a

Activation of heat shock proteins (Hsps) is critical to adaptation to low oxygen levels (hypoxia) and for enduring the oxidative stress of reoxygenation. Hsps are known to be regulated by heat shock factor (Hsf), but our results demonstrate an unexpected regulatory link between the oxygen-sensing and heat shock pathways. Hsf transcription is up-regulated during hypoxia due to direct binding by hypoxia-inducible factor-1 (HIF-1) to HIF-1 response elements in an Hsf intron. This increase in Hsf transcripts is necessary for full Hsp induction during hypoxia and reoxygenation. The HIF-1-dependent increase in Hsps has a functional impact, as reduced production of Hsps decreases viability of adult flies exposed to hypoxia and reoxygenation. Thus, HIF-1 control of Hsf transcriptional levels is a regulatory mechanism for sensitizing heat shock pathway activity in order to maximize production of protective Hsps. This cross-regulation represents a mechanism by which the low oxygen response pathway has assimilated complex new functions by regulating the key transcriptional activator of the heat shock pathway.In order to endure oxygen deprivation, most eukaryotes utilize a conserved set of cellular adaptations (1). Many of these changes are brought about by the activation of the transcription factor hypoxia-inducible factor-1 (HIF-1), 2 a heterodimeric complex composed of HIF-1␣ and HIF-1␤ subunits. When this complex is formed it binds to specific DNA enhancer sequences and regulates the activity of target genes. Both HIF-1␣ and HIF-1␤ are constitutively expressed in normal oxygen conditions (normoxia), but HIF-1␣ protein is quickly degraded before dimerization can occur with HIF-1␤ (2). Normoxic HIF-1␣ degradation is mediated by a series of hydroxylations and ubiquitinations that tag HIF-1␣ for disposal through the proteasome (3-6).The HIF-1 complex transcriptionally regulates a wide array of genes involved in anaerobic metabolism, growth, proliferation, angiogenesis, and cell death (7,8). This multifaceted control of cellular and organismal physiological pathways is exploited by solid tumors through the natural hypoxic environment caused by rapid growth or genetic alterations that stabilize HIF-1␣ (9). Overexpression or activation of HIF-1␣ is often seen in a wide array of cancers and is correlated with patient survival (10), and studies have shown that targeting the HIF-1 pathway is a promising means of cancer therapy (11,12). Thus, HIF-1 is a central regulator of normal and pathological changes in response to low oxygen.Although many genes that are up-regulated during hypoxia are known to be regulated by HIF-1, there are also diverse sets of genes up-regulated that have not been linked to the actions of HIF-1. Among these are the highly conserved heat shock proteins (Hsps) that are highly up-regulated during hypoxia but have not been linked to HIF-1 regulation (13). Hsps are known to act as cellular chaperones for proteins that are misfolded by cellular stresses (14). Heat shock factor (Hsf) was one of the first studied...

To study essential maternal gene requirements in the early C. elegans embryo, we have screened for temperature-sensitive, embryonic lethal mutations in an effort to bypass essential zygotic requirements for such genes during larval and adult germline development. With conditional alleles, multiple essential requirements can be examined by shifting at different times from the permissive temperature of 15°C to the restrictive temperature of 26°C. Here we describe 24 conditional mutations that affect 13 different loci and report the identity of the gene mutations responsible for the conditional lethality in 22 of the mutants. All but four are mis-sense mutations, with two mutations affecting splice sites, another creating an in-frame deletion, and one creating a premature stop codon. Almost all of the mis-sense mutations affect residues conserved in orthologs, and thus may be useful for engineering conditional mutations in other organisms. We find that 62% of the mutants display additional phenotypes when shifted to the restrictive temperature as L1 larvae, in addition to causing embryonic lethality after L4 upshifts. Remarkably, we also found that 13 out of the 24 mutations appear to be fast-acting, making them particularly useful for careful dissection of multiple essential requirements. Our findings highlight the value of C. elegans for identifying useful temperature-sensitive mutations in essential genes, and provide new insights into the requirements for some of the affected loci.

Total knee arthroplasty (TKA) is the most common and cost‐effective treatment for older adults with long‐standing osteoarthritis. Projections indicate that nearly 3.5 million older adults will undergo this procedure annually by the year 2030. Thus, understanding the factors that lead to optimal outcomes is of great clinical interest. In the majority of cases, tourniquet is applied during surgery to maintain a clear surgical field, however, there is debate as to whether this intervention is completely benign. In particular, muscle atrophy is a significant factor in preventing full functional recovery following surgery, and some evidence suggests that tourniquet application and the associated ischemia–reperfusion injury that results contributes to muscle atrophy. For this reason, we examined tissue level changes in muscle in TKA patients following surgery and found that there was a significant increase in cross‐sectional area of muscle fibers of all types. Furthermore, to detect changes not evident at the tissue level, we performed NextSeq analysis to assess the transcriptional landscape of quadriceps muscle cells following TKA with tourniquet and found 72 genes that were significantly upregulated. A large proportion of those genes regulate cell stress pathways, suggesting that muscle cells in our cohort of older adults were capable of mounting a significant response to cell stress. Furthermore, factors related to complement were upregulated, suggesting tourniquet may play a role in priming cells to ischemia reperfusion injury. Therefore, our analysis reveals potential harms of tourniquet during TKA, thus suggesting that surgeons should consider limiting its use.

Key pointsr Histamine is a primordial signalling molecule, capable of activating cells in an autocrine or paracrine fashion via specific cell surface receptors, in a variety of pathways that probably predate its more recent role in innate and adaptive immunity.r Although histamine is normally associated with pathological conditions or allergic and anaphylactic reactions, it may contribute beneficially to the normal changes that occur within skeletal muscle during the recovery from exercise.r We show that the human response to exercise includes an altered expression of thousands of protein-coding genes, and much of this response appears to be driven by histamine.r Histamine may be an important molecular transducer contributing to many of the adaptations that accompany chronic exercise training.Abstract Histamine is a primordial signalling molecule, capable of activating cells in an autocrine or paracrine fashion via specific cell surface receptors. In humans, aerobic exercise is followed by a post-exercise activation of histamine H 1 and H 2 receptors localized to the previously exercised muscle. This could trigger a broad range of cellular adaptations in response to exercise. Thus, we exploited RNA sequencing to explore the effects of H 1 and H 2 receptor blockade on the exercise transcriptome in human skeletal muscle tissue harvested from the vastus lateralis. We found that exercise exerts a profound influence on the human transcriptome, causing the differential expression of more than 3000 protein-coding genes. The influence of histamine blockade post-exercise was notable for 795 genes that were differentially expressed between the control and blockade condition, which represents >25% of the number responding to exercise. The broad histamine footprint on the human exercise transcriptome crosses many cellular functions, including inflammation, vascular function, metabolism, and cellular maintenance.