Cellular activities such as compound synthesis often require the transcriptional activation of an entire pathway; however, the molecular mechanisms underlying pathway activation have rarely been explained. Here, the cis regulatory architecture of the anthocyanin pathway genes targeted by the transcription factor (TF) complex including MYB, bHLH, and WDR was systematically analysed in one species and the findings extended to others. In Ipomoea purpurea, the IpMYB1-IpbHLH2-IpWDR1 (IpMBW) complex was found to be orthologous to the PAP1-GL3-TTG1 (AtPGT) complex of Arabidopsis thaliana, and interacted with a 7-bp MYB-recognizing element (MRE) and a 6-bp bHLH-recognizing element (BRE) at the proximal promoter region of the pathway genes. There was little transcription of the gene in the absence of the MRE or BRE. The cis elements identified experimentally converged on two syntaxes, ANCNNCC for MREs and CACN(A/C/T)(G/T) for BREs, and our bioinformatic analysis showed that these were present within anthocyanin gene promoters in at least 35 species, including both gymnosperms and angiosperms. For the anthocyanin pathway, IpMBW and AtPGT recognized the interspecific promoters of both early and later genes. In A. thaliana, the seed-specific TF complex (TT2, TT8, and TTG1) may regulate all the anthocyanin pathway genes, in addition to the proanthocyanidin-specific BAN. When multiple TF complexes in the anthocyanin pathway were compared, the cis architecture played a role larger than the TF complex in determining the variation in promoter activity. Collectively, a cis logic common to the pathway gene promoters was found, and this logic is essential for the trans factors to regulate the pathway.

Author Summary Candida is part of the gut microflora in healthy individuals, but can disseminate and cause systemic disease when the host's immune system is suppressed. Its ability to grow as yeast and hyphae in response to environmental cues is a major virulence attribute. Hyphal development requires temporary clearing of the transcription inhibitor Nrg1 upon activation of cAMP/PKA for initiation and promoter recruitment of the histone deacetylase Hda1 under reduced Tor1 signaling for maintenance. Here, we show that, during hyphal initiation when Nrg1 is gone, expression of the GATA family transcription factor Brg1 is activated under reduced Tor1 signaling. Accumulated Brg1 recruits Hda1 to hyphal promoters to reposition nucleosomes, leading to obstruction of Nrg1 binding sites and sustained hyphal development. The nucleosome repositioning during the yeast-hyphal transition provides a mechanism for temporal integration of extracellular signals and cell-fate specification. The hypha-specific transcription factor Ume6 functions after Brg1 in this succession of feed-forward regulation of hyphal development. Since misregulation of either Nrg1 or Ume6 causes altered virulence, and Brg1 regulates both Nrg1 accessibility and Ume6 transcription, our findings should provide a better understanding of how Candida controls its morphological program in different host niches to exist as a commensal and a pathogen.

American wild‐rice (Zizania palustris var. palustris) has served as a staple for indigenous North Americans for thousands of years, but has had significant habitat losses in recent centuries. We investigated genetic variability among 17 wild‐rice populations in northern Wisconsin using 13 isozyme markers. We then compared these genetic patterns to differences in habitat and population characteristics and phenotypic variation in plant growth and reproduction across sites. Wild‐rice's mean genetic diversity (0.15) is moderate compared to wind‐pollinated outcrossers but lower than the mean (0.20) reported for the Poaceae. Estimated inbreeding coefficients within populations (f) average 0.12 but vary greatly among the populations (from −0.44–0.52), suggesting heterogeneous population histories. Larger populations in larger lakes express higher levels of genetic variability and smaller inbreeding coefficients than smaller or more isolated populations. The number of panicles per plant is also higher in populations with greater genetic variability. Estimated genetic differentiation among the 17 populations (F_ST) was high (0.30), suggesting limited gene flow among drainages. Wild‐rice population size and degree of isolation have opposing effects on its genetic variability, and plant performance is positively associated with genetic variability.

Background: Transcription factor (TF) binding sites (cis element) play a central role in gene regulation, and eukaryotic organisms frequently adapt a combinatorial regulation to render sophisticated local gene expression patterns. Knowing the precise cis element on a distal promoter is a prerequisite for studying a typical transcription process; however, identifications of cis elements have lagged behind those of their associated trans acting TFs due to technical difficulties. Consequently, gene regulations via combinatorial TFs, as widely observed across biological processes, have remained vague in many cases. Results: We present here a valid strategy for identifying cis elements in combinatorial TF regulations. It consists of bioinformatic searches of available databases to generate candidate cis elements and tests of the candidates using improved experimental assays. Taking the MYB and the bHLH that collaboratively regulate the anthocyanin pathway genes as examples, we demonstrate how candidate cis motifs for the TFs are found on multi-specific promoters of chalcone synthase (CHS) genes, and how to experimentally test the candidate sites by designing DNA fragments hosting the candidate motifs based on a known promoter (us1 allele of Ipomoea purpurea CHS-D in our case) and applying site-mutagenesis at the motifs. It was shown that TF-DNA interactions could be unambiguously analyzed by assays of electrophoretic mobility shift (EMSA) and dual-luciferase transient expressions, and the resulting evidence precisely delineated a cis element. The cis element for R2R3 MYBs including Ipomoea MYB1 and Magnolia MYB1, for instance, was found to be ANCNACC, and that for bHLHs (exemplified by Ipomoea bHLH2 and petunia AN1) was CACNNG. A re-analysis was conducted on previously reported promoter segments recognized by maize C1 and apple MYB10, which indicated that cis elements similar to ANCNACC were indeed present on these segments, and tested positive for their bindings to Ipomoea MYB1. Conclusion: Identification of cis elements in combinatorial regulation is now feasible with the strategy outlined. The working pipeline integrates the existing databases with experimental techniques, providing an open framework for precisely identifying cis elements. This strategy is widely applicable to various biological systems, and may enhance future analyses on gene regulation.

Background: Asian rice (Oryza sativa L.) has been a model plant but its cultivation history is inadequately understood, and its origin still under debate. Several enigmas remain, including how this annual crop shifted its growth habit from its perennial ancestor, O. rufipogon, why genetic divergence between indica and japonica appears older than the history of human domestication, and why some domestication genes do not show signals of introgression between subgroups. Addressing these issues may benefit both basic research and rice breeding. Results: Gene genealogy-based mutation (GGM) analysis shows that history of Asian rice is divided into two phases (Phase I and II) of about equal lengths. Mutations occurred earlier than the partition of indica and japonica to Os genome mark Phase-I period. We diagnosed 91 such mutations among 101 genes sampled across 12 chromosomes of Asian rice and its wild relatives. Positive selection, detected more at 5′ regions than at coding regions of some of the genes, involved 22 loci (e.g., An-1, SH4, Rc, Hd3a, GL3.2, OsMYB3, OsDFR, and OsMYB15), which affected traits from easy harvesting, grain color, flowering time, productivity, to likely taste and tolerance. Phase-I mutations of OsMYB3, OsHd3a and OsDFR were experimentally tested and all caused enhanced functions of the genes in vivo. Phase-II period features separate cultivations, lineage-specific selection, and expanded domestication to more genes. Further genomic analysis, along with phenotypic comparisons, indicates that O. sativa is hybrid progeny of O. rufipogon and O. nivara, inherited slightly more genes of O. rufipogon. Congruently, modern alleles of the sampled genes are approximately 6% ancient, 38% uni-specific, 40% bi-specific (mixed), and 15% new after accumulating significant mutations. Results of sequencing surveys across modern cultivars/landraces indicate locus-specific usages of various alleles while confirming the associated mutations. Conclusions: Asian rice was initially domesticated as one crop and later separate selection mediated by human resulted in its major subgroups. This history and the hybrid origin well explain previous puzzles. Positive selection, particularly in 5′ regions, was the major force underlying trait domestication. Locus-specific domestication can be characterized and the result may facilitate breeders in developing better rice varieties in future.