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Species that repeatedly evolve phenotypic clines across environmental gradients have been highlighted as ideal systems for characterizing the genomic basis of local environmental adaptation. However, few studies have assessed the importance of observed phenotypic clines for local adaptation: conspicuous traits that vary clinally may not necessarily be the most critical in determining local fitness. The present study was designed to fill this gap, using a plant species characterized by repeatedly evolved adaptive phenotypic clines. White clover is naturally polymorphic for its chemical defence cyanogenesis (HCN release with tissue damage); climate‐associated cyanogenesis clines have evolved throughout its native and introduced range worldwide. We performed landscape genomic analyses on 415 wild genotypes from 43 locations spanning much of the North American species range to assess the relative importance of cyanogenesis loci vs. other genomic factors in local climatic adaptation. We find clear evidence of local adaptation, with temperature‐related climatic variables best describing genome‐wide differentiation between sampling locations. The same climatic variables are also strongly correlated with cyanogenesis frequencies and gene copy number variations (CNVs) at cyanogenesis loci. However, landscape genomic analyses indicate no significant contribution of cyanogenesis loci to local adaptation. Instead, several genomic regions containing promising candidate genes for plant response to seasonal cues are identified — some of which are shared with previously identified QTLs for locally adaptive fitness traits in North American white clover. Our findings suggest that local adaptation in white clover is likely determined primarily by genes controlling the timing of growth and flowering in response to local seasonal cues. More generally, this work suggests that caution is warranted when considering the importance of conspicuous phenotypic clines as primary determinants of local adaptation. [ABSTRACT FROM AUTHOR]

Heterotrimeric G protein signaling is essential for normal hyphal growth in the filamentous fungus Neurospora crassa. We have previously demonstrated that the non-receptor guanine nucleotide exchange factor RIC8 acts upstream of the Gα proteins GNA-1 and GNA-3 to regulate hyphal extension. Here we demonstrate that regulation of hyphal extension results at least in part, from an important role in control of asexual spore (conidia) germination. Loss of GNA-3 leads to a drastic reduction in conidial germination, which is exacerbated in the absence of GNA-1. Mutation of RIC8 leads to a reduction in germination similar to that in the Δgna-1, Δgna-3 double mutant, suggesting that RIC8 regulates conidial germination through both GNA-1 and GNA-3. Support for a more significant role for GNA-3 is indicated by the observation that expression of a GTPase-deficient, constitutively active gna-3 allele in the Δric8 mutant leads to a significant increase in conidial germination. Localization of the three Gα proteins during conidial germination was probed through analysis of cells expressing fluorescently tagged proteins. Functional TagRFP fusions of each of the three Gα subunits were constructed through insertion of TagRFP in a conserved loop region of the Gα subunits. The results demonstrated that GNA-1 localizes to the plasma membrane and vacuoles, and also to septa throughout conidial germination. GNA-2 and GNA-3 localize to both the plasma membrane and vacuoles during early germination, but are then found in intracellular vacuoles later during hyphal outgrowth.

Local adaptation is common in plants, yet characterization of its underlying genetic basis is rare in herbaceous perennials. Moreover, while many plant species exhibit intraspecific chemical defence polymorphisms, their importance for local adaptation remains poorly understood. We examined the genetic architecture of local adaptation in a perennial, obligately‐outcrossing herbaceous legume, white clover (Trifolium repens). This widespread species displays a well‐studied chemical defence polymorphism for cyanogenesis (HCN release following tissue damage) and has evolved climate‐associated cyanogenesis clines throughout its range. Two biparental F2 mapping populations, derived from three parents collected in environments spanning the U.S. latitudinal species range (Duluth, MN, St. Louis, MO and Gainesville, FL), were grown in triplicate for two years in reciprocal common garden experiments in the parental environments (6,012 total plants). Vegetative growth and reproductive fitness traits displayed trade‐offs across reciprocal environments, indicating local adaptation. Genetic mapping of fitness traits revealed a genetic architecture characterized by allelic trade‐offs between environments, with 100% and 80% of fitness QTL in the two mapping populations showing significant QTL×E interactions, consistent with antagonistic pleiotropy. Across the genome there were three hotspots of QTL colocalization. Unexpectedly, we found little evidence that the cyanogenesis polymorphism contributes to local adaptation. Instead, divergent life history strategies in reciprocal environments were major fitness determinants: selection favoured early investment in flowering at the cost of multiyear survival in the southernmost site versus delayed flowering and multiyear persistence in the northern environments. Our findings demonstrate that multilocus genetic trade‐offs contribute to contrasting life history characteristics that allow for local adaptation in this outcrossing herbaceous perennial. see also the Perspective by Jill T. Anderson [ABSTRACT FROM AUTHOR]

The article analyzes heterotrimeric G protein in filamentous fungi by identifying G protein-coupled receptor genes. The sensing of nutrients and host signals of G proteins is useful for pathogenesis.

G protein-coupled receptors (GPCRs) are the largest family of human proteins. They have a common structure and, signaling through a much smaller set of G proteins, arrestins, and effectors, activate downstream pathways that often modulate hallmark mechanisms of cancer. Because there are many more GPCRs than effectors, mutations in different receptors could perturb signaling similarly so as to favor a tumor. We hypothesized that somatic mutations in tumor samples may not be enriched within a single gene but rather that cognate mutations with similar effects on GPCR function are distributed across many receptors. To test this possibility, we systematically aggregated somatic cancer mutations across class A GPCRs and found a nonrandom distribution of positions with variant amino acid residues. Individual cancer types were enriched for highly impactful, recurrent mutations at selected cognate positions of known functional motifs. We also discovered that no single receptor drives this pattern, but rather multiple receptors contain amino acid substitutions at a few cognate positions. Phenotypic characterization suggests these mutations induce perturbation of G protein activation and/or β-arrestin recruitment. These data suggest that recurrent impactful oncogenic mutations perturb different GPCRs to subvert signaling and promote tumor growth or survival. The possibility that multiple different GPCRs could moonlight as drivers or enablers of a given cancer through mutations located at cognate positions across GPCR paralogs opens a window into cancer mechanisms and potential approaches to therapeutics. [ABSTRACT FROM AUTHOR]

Beta adrenergic receptors (βARs) are G-protein-coupled receptors essential for physiological responses to the hormones/neurotransmitters epinephrine and norepinephrine which are found in the nervous system and throughout the body. They are the targets of numerous widely used drugs, especially in the case of the most extensively studied βAR, β2AR, whose ligands are used for asthma and cardiovascular disease. βARs signal through Gαs G-proteins and via activation of adenylyl cyclase and cAMP-dependent protein kinase, but some alternative downstream pathways have also been proposed that could be important for understanding normal physiological functioning of βAR signaling and its disruption in disease. Using fluorescence-based Ca2+ flux assays combined with pharmacology and gene knock-out methods, we discovered a previously unrecognized endogenous pathway in HEK-293 cells whereby β2AR activation leads to robust Ca2+ mobilization from intracellular stores via activation of phospholipase C and opening of inositol trisphosphate (InsP3) receptors. This pathway did not involve cAMP, Gαs, or Gαi or the participation of the other members of the canonical β2AR signaling cascade and, therefore, constitutes a novel signaling mechanism for this receptor. This newly uncovered mechanism for Ca2+mobilization byβ2ARhas broad implications for adrenergic signaling, cross-talk with other signaling pathways, and the effects of βAR-directed drugs. [ABSTRACT FROM AUTHOR]

Seed oil melting point is an adaptive, quantitative trait determined by the relative proportions of the fatty acids that compose the oil. Micro- and macro-evolutionary evidence suggests selection has changed the melting point of seed oils to covary with germination temperatures because of a trade-off between total energy stores and the rate of energy acquisition during germination under competition. The seed oil compositions of 391 natural accessions of Arabidopsis thaliana, grown under common-garden conditions, were used to assess whether seed oil melting point within a species varied with germination temperature. In support of the adaptive explanation, long-term monthly spring and fall field temperatures of the accession collection sites significantly predicted their seed oil melting points. In addition, a genomewide association study (GWAS) was performed to determine which genes were most likely responsible for the natural variation in seed oil melting point. The GWAS found a single highly significant association within the coding region of FAD2, which encodes a fatty acid desaturase central to the oil biosynthesis pathway. In a separate analysis of 15 a priori oil synthesis candidate genes, 2 (FAD2 and FATB) were located near significant SNPs associated with seed oil melting point. These results comport with others' molecular work showing that lines with alterations in these genes affect seed oil melting point as expected. Our results suggest natural selection has acted on a small number of loci to alter a quantitative trait in response to local environmental conditions. [ABSTRACT FROM AUTHOR]

The renewable source of highly reduced carbon provided by plant triacylglycerols (TAGs) fills an ever increasing demand for food, biodiesel, and industrial chemicals. Each of these uses requires different compositions of fatty acid proportions in seed oils. Identifying the genes responsible for variation in seed oil composition in nature provides targets for bioengineering fatty acid proportions optimized for various industrial and nutrition goals. Here, we characterized the seed oil composition of 391 world-wide, wild accessions of Arabidopsis thaliana, and performed a genome-wide association study (GWAS) of the 9 major fatty acids in the seed oil and 4 composite measures of the fatty acids. Four to 19 regions of interest were associated with the seed oil composition traits. Thirty-four of the genes in these regions are involved in lipid metabolism or transport, with 14 specific to fatty acid synthesis or breakdown. Eight of the genes encode transcription factors. We have identified genes significantly associated with variation in fatty acid proportions that can be used as a resource across the Brassicaceae. Two-thirds of the regions identified contain candidate genes that have never been implicated in lipid metabolism and represent potential new targets for bioengineering. [ABSTRACT FROM AUTHOR]

Here we characterize the relationship between the PRE-2 pheromone receptor and its ligand, CCG-4, and the general requirements for receptors, pheromones, G proteins, and mating type genes during fusion of opposite mating-type cells and sexual sporulation in the multicellular fungus Neurospora crassa. PRE-2 is highly expressed in mat a cells and is localized in male and female reproductive structures. Δpre-2 mat a females do not respond chemotropically to mat A males (conidia) or form mature fruiting bodies (perithecia) or meiotic progeny (ascospores). Strains with swapped identity due to heterologous expression of pre-2 or ccg-4 behave normally in crosses with opposite mating-type strains. Coexpression of pre-2 and ccg-4 in the mat A background leads to self-attraction and development of barren perithecia without ascospores. Further perithecial development is achieved by inactivation of Sad-1, a gene required for meiotic gene silencing. Findings from studies involving forced heterokaryons of opposite mating-type strains show that presence of one receptor and its compatible pheromone is necessary and sufficient for perithecial development and ascospore production. Taken together, the results demonstrate that although receptors and pheromones control sexual identity, the mating-type genes (mat A and mat a) must be in two different nuclei to allow meiosis and sexual sporulation to occur. [ABSTRACT FROM AUTHOR]

Beta adrenergic receptors (βARs) are G-protein-coupled receptors essential for physiological responses to the hormones/neurotransmitters epinephrine and norepinephrine which are found in the nervous system and throughout the body. They are the targets of numerous widely used drugs, especially in the case of the most extensively studied βAR, β 2 AR, whose ligands are used for asthma and cardiovascular disease. βARs signal through Gα s G-proteins and via activation of adenylyl cyclase and cAMP-dependent protein kinase, but some alternative downstream pathways have also been proposed that could be important for understanding normal physiological functioning of βAR signaling and its disruption in disease. Using fluorescence-based Ca 2+ flux assays combined with pharmacology and gene knock-out methods, we discovered a previously unrecognized endogenous pathway in HEK-293 cells whereby β 2 AR activation leads to robust Ca 2+ mobilization from intracellular stores via activation of phospholipase C and opening of inositol trisphosphate (InsP 3 ) receptors. This pathway did not involve cAMP, Gα s , or Gα i or the participation of the other members of the canonical β 2 AR signaling cascade and, therefore, constitutes a novel signaling mechanism for this receptor. This newly uncovered mechanism for Ca 2+ mobilization by β 2 AR has broad implications for adrenergic signaling, cross-talk with other signaling pathways, and the effects of βAR-directed drugs., Competing Interests: The authors declare that they have no conflicts of interest with the contents of this article. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health., (© 2017 by The American Society for Biochemistry and Molecular Biology, Inc.)