Homeostatic control of blood glucose requires different physiological responses in the fasting and post-prandial states. We reasoned that glucose measurements under non-standardised conditions (random glucose, RG) may capture diverse glucoregulatory processes more effectively than previous genome-wide association studies (GWAS) of fasting glycaemia or after standardised glucose loads. Through GWAS meta-analysis of RG in 493,036 individuals without diabetes of diverse ethnicities we identified 128 associated loci represented by 162 distinct signals, including 14 with sex-dimorphic effects, 9 discovered through trans-ethnic analysis, and 70 novel signals for glycaemic traits. Novel RG loci were particularly enriched in expression in the ileum and colon, indicating a prominent role for the gastrointestinal tract in the control of blood glucose. Functional studies and molecular dynamics simulations of coding variants of GLP1R, a well-established type 2 diabetes treatment target, provided a genetic framework for optimal selection of GLP-1R agonist therapy. We also provided new evidence from Mendelian randomisation that lung function is modulated by blood glucose and that pulmonary dysfunction is a diabetes complication. Thus, our approach based on RG GWAS provided wide-ranging insights into the biology of glucose regulation, diabetes complications and the potential for treatment stratification.

AbbreviationscAMP Cyclic adenosine monophosphate DERET Diffusion-enhanced resonance energy transfer DPP-4 Dipeptidyl dipeptidase-4 GCG Glucagon GCGR Glucagon receptor GIP Glucose-dependent insulinotropic polypeptide GIPR Glucose-dependent insulinotropic polypeptide receptor GLP-1 Glucagon-like peptide-1 GLP-1R Glucagon-like peptide-1 receptor NEP24.11 Neutral endopeptidase T2D Type 2 diabetes TR-FRET Time-resolved Förster resonance energy transfer AbstractReceptors for the peptide hormones glucagon-like peptide-1 (GLP-1), glucose-dependent insulinotropic polypeptide (GIP) and glucagon (GCG) are important regulators of insulin secretion and energy metabolism. Here we sought to investigate how signal bias between cyclic AMP and βarrestin-2 recruitment can modulate the effects of prolonged agonist stimulation at each of these receptors. We generated analogues of GLP-1, GCG and GIP which in some cases showed selective reduction in β-arrestin-2 recruitment versus cAMP signalling compared to the parent peptide. Despite reduced acute signalling potency and/or efficacy, some biased GLP-1 and GIP analogues increased maximal sustained insulin secretion, although only at high agonist concentrations. Biased GCG analogues did not affect maximal insulin release from beta cells, or glucose output in hepatocytes.

In eukaryotes, splice sites define the introns of pre-mRNAs and must be recognized and excised with nucleotide precision by the spliceosome to make the correct mRNA product. In one of the earliest steps of spliceosome assembly, the U1 small nuclear ribonucleoprotein (snRNP) recognizes the 5' splice site (5' SS) through a combination of base pairing, protein-RNA contacts, and interactions with other splicing factors. Previous studies investigating the mechanisms of 5' SS recognition have largely been done in vivo or in cellular extracts where the U1/5' SS interaction is difficult to deconvolute from the effects of trans-acting factors or RNA structure. In this work we used co-localization single-molecule spectroscopy (CoSMoS) to elucidate the pathway of 5' SS selection by purified yeast U1 snRNP. We determined that U1 reversibly selects 5' SS in a sequence-dependent, two-step mechanism. A kinetic selection scheme enforces pairing at particular positions rather than overall duplex stability to achieve long-lived U1 binding. Our results provide a kinetic basis for how U1 may rapidly surveil nascent transcripts for 5' SS and preferentially accumulate at these sequences rather than on close cognates.