2020 design and methods of the Population Assessment of Tobacco and Health (PATH) study during the COVID-19 pandemic.

The Population Assessment of Tobacco and Health (PATH) Study is a nationally representative, longitudinal study of the US population on tobacco use and its effects on health, collecting data annually since 2013. The COVID-19 pandemic interrupted in-person survey data collections around the world. In the USA, this included a PATH Study data collection focused on youth (13-17) and young adults (18-19) as well as other US surveys on tobacco use. Given that it was necessary to pause data collection and considering that tobacco-use behaviours could be expected to change along with pandemic-related changes in the social environment, the original design for the 2020 PATH Study data collection for youth and young adults was modified. Also, the PATH Study Adult Telephone Survey was developed to address the need for adult tobacco use monitoring in this unprecedented time. This article describes the modifications made to the 2020 PATH Study design and protocol to provide nationally representative data for youth and adults after the onset of the COVID-19 pandemic as well as the implications of these modifications for researchers.

Serine arginine splicing factor 3 is involved in enhanced splicing of glucose-6-phosphate dehydrogenase RNA in response to nutrients and hormones in liver.

Expression of G6PD is controlled by changes in the degree of splicing of the G6PD mRNA in response to nutrients in the diet. This regulation involves an exonic splicing enhancer (ESE) in exon 12 of the mRNA. Using the G6PD model, we demonstrate that nutrients and hormones control the activity of serine-arginine-rich (SR) proteins, a family of splicing co-activators, and thereby regulate the splicing of G6PD mRNA. In primary rat hepatocyte cultures, insulin increased the amount of phosphorylated SR proteins, and this effect was counteracted by arachidonic acid. The results of RNA affinity analysis with nuclear extracts from intact liver demonstrated that the SR splicing factor proteins SRSF3 and SRSF4 bound to the G6PD ESE. Consequently, siRNA-mediated depletion of SRSF3, but not SRSF4, in liver cells inhibited accumulation of both mRNA expressed from a minigene containing exon 12 and the endogenous G6PD mRNA. Consistent with the functional role of SRSF3 in regulating splicing, SRSF3 was observed to bind to the ESE in both intact cells and in animals using RNA immunoprecipitation analysis. Furthermore, refeeding significantly increased the binding of SRSF3 coincident with increased splicing and expression of G6PD. Together, these data establish that nutritional regulation of SRSF3 activity is involved in the differential splicing of the G6PD transcript in response to nutrients. Nutritional regulation of other SR proteins presents a regulatory mechanism that could cause widespread changes in mRNA splicing. Nutrients are therefore novel regulators of mRNA splicing.

CM156, a sigma receptor ligand, reverses cocaine-induced place conditioning and transcriptional responses in the brain.

Repeated exposure to cocaine induces neuroadaptations which contribute to the rewarding properties of cocaine. Using cocaine-induced conditioned place preference (CPP) as an animal model of reward, earlier studies have shown that sigma (σ) receptor ligands can attenuate the acquisition, expression and reactivation of CPP. However, the underlying molecular mechanisms that are associated with these changes are not yet understood. In the present study, CM156, a novel antagonist with high selectivity and affinity for σ receptors was used to attenuate the expression of cocaine-induced CPP in mice. Immediately following the behavioral evaluations, mouse brain tissues were collected and alterations in gene expression in half brain samples were profiled by cDNA microarray analysis. Microarray data was analyzed by three distinct normalization methods and four genes were consistently found to be upregulated by cocaine when compared to saline controls. Each of these gene changes were found by more than one normalization method to be reversed by at least one dose of CM156. Quantitative real time PCR confirmed that a single administration of CM156 was able to reverse the cocaine-induced increases in three of these four genes: metastasis associated lung adenocarcinoma transcript 1 (malat1), tyrosine 3-monooxygenase/tryptophan 5-monooxygenase activation protein (ywhaz), and transthyretin (ttr). These genes are involved in processes related to neuroplasticity and RNA editing. The data presented herein provides evidence that pharmacological intervention with a putative σ receptor antagonist reverses alterations in gene expression that are associated with cocaine-induced reward.

Arachidonic acid inhibits the insulin induction of glucose-6-phosphate dehydrogenase via p38 MAP kinase.

Polyunsaturated fatty acids are potent inhibitors of lipogenic gene expression in liver. The lipogenic enzyme glucose-6-phosphate dehydrogenase (G6PD) is unique in this gene family, in that fatty acids inhibit at a post-transcriptional step. In this study, we have provided evidence for a signaling pathway for the arachidonic acid inhibition of G6PD mRNA abundance. Arachidonic acid decreases the insulin induction of G6PD expression; by itself, arachidonic acid does not inhibit basal G6PD mRNA accumulation. The insulin stimulation of G6PD involves the phosphoinositide 3-kinase (PI 3-kinase) pathway (Wagle, A., Jivraj, S., Garlock, G. L., and Stapleton, S. R. (1998) J. Biol. Chem. 273, 14968-14974). Incubation of hepatocytes with arachidonic acid blocks the activation of PI 3-kinase by insulin as observed by a decrease in Ser(473) phosphorylation of Akt, the downstream effector of PI 3-kinase. The decrease in PI 3-kinase activity was associated with an increase in Ser(307) phosphorylation of IRS-1. Western analysis demonstrated increased phosphorylation of p38 mitogen-activated protein kinase (MAPK) in arachidonic acid-treated cells, whereas extracellular signal-regulated kinase and c-Jun NH(2)-terminal kinase activity was not changed. Incubating the hepatocytes with the p38 MAPK inhibitor, SB203580, blocked the arachidonic acid inhibition of G6PD mRNA accumulation. Furthermore, SB203580 decreased the arachidonic acid-mediated Ser(307) phosphorylation of IRS-1 and rescued Akt activation that was otherwise decreased by arachidonic acid. Thus, arachidonic acid inhibits the insulin stimulation of G6PD mRNA accumulation by stimulating the p38 MAPK pathway, thereby inhibiting insulin signal transduction.