Whole genome expression profiles of yeast RNA polymerase II core subunit, Rpb4, in stress and nonstress conditions

Organisms respond to environmental stress by adopting changes in gene expression at the transcriptional level. Rpb4, a nonessential subunit of the core RNA polymerase II has been proposed to play a role in non-stress-specific transcription and in the regulation of stress response in yeast. We find that in addition to the temperature sensitivity of the null mutant of Rpb4, diploid null mutants are also compromised in sporulation and show morphological changes associated with nitrogen starvation. Using whole genome expression analysis, we report here the effects of Rpb4 on expression of genes during normal growth and following heat shock and nutritional starvation. Our analysis shows that Rpb4 affects expression of a small yet significant fraction of the genome in both stress and normal conditions. We found that genes involved in galactose metabolism were dependent on the presence of Rpb4 irrespective of the environmental condition. Rpb4 was also found to affect the expression of several other genes specifically in conditions of nutritional starvation. The general defect in the absence of Rpb4 is in the expression of metabolic genes, especially those involved in carbon metabolism and energy generation. We report that various stresses are affected byRPB4 and that on overexpression the stress-specific activators can partially rescue the corresponding defects

Metabolomics Investigation Reveals Metabolite Mediators Associated with Acute Lung Injury and Repair in a Murine Model of Influenza Pneumonia

Influenza virus infection (IVI) can cause primary viral pneumonia, which may progress to acute lung injury (ALI) and respiratory failure with a potentially fatal outcome. At present, the interactions between host and influenza virus at molecular levels and the underlying mechanisms that give rise to IVI-induced ALI are poorly understood. We conducted a comprehensive mass spectrometry-based metabolic profiling of serum, lung tissue and bronchoalveolar lavage fluid (BALF) from a non-lethal mouse model with influenza A virus at 0, 6, 10, 14, 21 and 28 days post infection (dpi), representing the major stages of IVI. Distinct metabolite signatures were observed in mice sera, lung tissues and BALF, indicating the molecular differences between systematic and localized host responses to IVI. More than 100 differential metabolites were captured in mice sera, lung tissues and BALF, including purines, pyrimidines, acylcarnitines, fatty acids, amino acids, glucocorticoids, sphingolipids, phospholipids, etc. Many of these metabolites belonged to pulmonary surfactants, indicating IVI-induced aberrations of the pulmonary surfactant system might play an important role in the etiology of respiratory failure and repair. Our findings revealed dynamic host responses to IVI and various metabolic pathways linked to disease progression, and provided mechanistic insights into IVI-induced ALI and repair process.National Science Foundation (U.S.)Singapore-MIT Alliance for Research and Technology (SMART

Molecular Analysis of Serum and Bronchoalveolar Lavage in a Mouse Model of Influenza Reveals Markers of Disease Severity That Can Be Clinically Useful in Humans

Background: Management of influenza, a major contributor to the worldwide disease burden, is complicated by lack of reliable methods for early identification of susceptible individuals. Identification of molecular markers that can augment existing diagnostic tools for prediction of severity can be expected to greatly improve disease management capabilities. Methodology/Principal Findings: We have analyzed cytokines, proteome flux and protein adducts in bronchoalveolar lavage (BAL) and sera from mice infected with influenza A virus (PR8 strain) using a previously established non-lethal model of influenza infection. Through detailed cytokine and protein adduct measurements of murine BAL, we first established the temporal profile of innate and adaptive responses as well as macrophage and neutrophil activities in response to influenza infection. A similar analysis was also performed with sera from a longitudinal cohort of influenza patients. We then used an iTRAQ-based, comparative serum proteome analysis to catalog the proteome flux in the murine BAL during the stages correlating with “peak viremia,” “inflammatory damage,” as well as the “recovery phase.” In addition to activation of acute phase responses, a distinct class of lung proteins including surfactant proteins was found to be depleted from the BAL coincident with their “appearance” in the serum, presumably due to leakage of the protein following loss of the integrity of the lung/epithelial barrier. Serum levels of at least two of these proteins were elevated in influenza patients during the febrile phase of infection compared to healthy controls or to the same patients at convalescence. Conclusions/Significance: The findings from this study provide a molecular description of disease progression in a mouse model of influenza and demonstrate its potential for translation into a novel class of markers for measurement of acute lung injury and improved case management.Singapore. National Research FoundationSingapore-MIT Alliance for Research and Technology (ID-IRG research program

Serum Metabolome and Lipidome Changes in Adult Patients with Primary Dengue Infection

10.1371/journal.pntd.0002373PLoS Neglected Tropical Diseases78

Serum Proteome and Cytokine Analysis in a Longitudinal Cohort of Adults with Primary Dengue Infection Reveals Predictive Markers of DHF

10.1371/journal.pntd.0001887PLoS Neglected Tropical Diseases611

Induced Hsp70 is in small, cytoplasmic complexes in a cell culture model of renal ischemia: a comparative study with heat shock

A number of clinical conditions are known to result in the induction of heat shock proteins, but detailed studies on stress response have focused mostly on heat shock as a model. We have analyzed the induction and intracellular distribution of heat shock proteins in a reversible adenosine triphosphate (ATP) depletion model of renal ischemia. Two Hsp70 homologues, Hsp70 in the cytoplasm and BiP in the endoplasmic reticulum (ER) lumen, were found significantly induced during the recovery phase of ATP depletion. Other members of the heat shock protein family, such as Hsp90, constitutive Hsc70, and a related protein Hop60, were not induced. The induction of stress proteins on ATP depletion differed from that after heat shock in the kinds of proteins elaborated, their induction kinetics, and their intracellular distributions. Biochemical fractionation and indirect immunofluorescence experiments indicated that Hsp70 was predominantly cytoplasmic in the recovery phase of ischemia-like stress. Velocity sedimentation on sucrose gradients showed that induced Hsp70 sedimented as small, soluble complexes, ranging in size from 4S(20,w) to 8S(20,w). The results suggest a role for induced Hsp70 that may be different from one of protecting aggregated proteins as under heat shock and emphasize the need for their characterization in other clinical conditions that result in stress response

Stress protein flux during recovery from simulated ischemia: Induced heat shock protein 70 confers cytoprotection by suppressing JNK activation and inhibiting apoptotic cell death

Multiple stress proteins are recruited in response to stress in living cells. There are limited reports in the literature analyzing multiple stress protein shifts and their functional consequences on stress response. Using two-dimensional electrophoresis we have analyzed shifts in stress protein profiles in response to energy deprivation as a model of ischemic injury to kidneys. A group of chaperones and stress-induced mitogen activated protein (MAP) kinases were analyzed. In addition to examining stress protein induction and phosphorylation we have also examined the mechanism of cytoprotection by heat shock protein 70 (Hsp70). Our results show that, of the different stress proteins examined, only binding protein (BiP) and Hsp70 were significantly induced upon energy deprivation. Other stress proteins, including Hsp27, calnexin, Hsp90 and ERp57 showed alterations in their phosphorylation profiles. Three different MAP kinases, namely p38, extracellular signal regulated kisase and c-jun N-terminal kinase (JNK) were activated in response to energy deprivation. While JNK activation was linked to apoptosis, activated-p38 was involved in phosphorylation of Hsp27. Study of inhibitors of Hsp70 induction or pre-induction of Hsp70 indicated that induced Hsp70 was involved in the suppression of JNK activation thereby inhibiting apoptotic cell death. Our results provide important insights into the flux in stress protein profiles in response to simulated ischemia and highlight the antiapoptotic, cytoprotective mechanism of Hsp70 action

Heat shock protein 70 as a biomarker of heat stress in a simulated hot cockpit

Background: Fighter pilots are frequently exposed to high temperatures during high-speed low-level flight. Heat strain can result in temporary impairment of cognitive functions and when severe, loss of consciousness and consequent loss of life and equipment. Induction of stress proteins is a highly conserved stress response mechanism from bacteria to humans. induced stress protein levels are known to be cytoprotective and have been correlated with stress tolerance. Although many studies on the heat shock response mechanisms have been performed in cell culture and animal model systems, there is very limited information on stress protein induction in human subjects. Hypothesis: Heat shock proteins (Hsp), especially Hsp70, may be induced in human subjects exposed to high temperatures in a hot cockpit designed to simulate heat stress experienced in low flying sorties. Methods: Six healthy volunteers were subjected to heat stress at 55degreesC in a high temperature cockpit simulator for a period of 1 h at 30% humidity. Physiological parameters such as oral and skin temperatures, heart rate, and sweat rate were monitored regularly during this time. The level of Hsp70 in leukocytes was examined before and after the heat exposure in each subject. Conclusions: Hsp70 was found to be significantly induced in all the six subjects exposed to heat stress. The level of induced Hsp70 appears to correlate with other strain indicators such as accumulative circulatory strain and Craig's modified index. The usefulness of Hsp70 as a molecular marker of heat stress in humans is discussed