Ramularia leaf spot disease (RLS), caused by the ascomycete fungus Ramularia collo-cygni, has emerged as a major economic disease of barley. No substantial resistance has been identified, so far, among barley genotypes and, based on the epidemiology of the disease, a quantitative genetic determinacy of RLS has been suggested. The relative contributions of barley and R. collo-cygni genetics to disease infection and epidemiology are practically unknown. Here, we present an integrated genome-wide analysis of host and pathogen transcriptome landscapes identified in a sensitive barley cultivar following infection by an aggressive R. collo-cygni isolate. We compared transcriptional responses in the infected and noninfected leaf samples in order to identify which molecular events are associated with RLS symptom development. We found a large proportion of R. collo-cygni genes to be expressed in planta and that many were also closely associated with the infection stage. The transition from surface to apoplastic colonization was associated with downregulation of cell wall–degrading genes and upregulation of nutrient uptake and resistance to oxidative stresses. Interestingly, the production of secondary metabolites was dynamically regulated within the fungus, indicating that R. collo-cygni produces a diverse panel of toxic compounds according to the infection stage. A defense response against R. collo-cygni was identified in barley at the early, asymptomatic infection and colonization stages. We found activation of ethylene signaling, jasmonic acid signaling, and phenylpropanoid and flavonoid pathways to be highly induced, indicative of a classical response to necrotrophic pathogens. Disease development was found to be associated with gene expression patterns similar to those found at the onset of leaf senescence, when nutrients, possibly, are used by the infecting fungus. These analyses, combining both barley and R. collo-cygni transcript profiles, demonstrate the activation of complex transcriptional programs in both organisms.

The ginkgo tree (Ginkgo biloba) is considered a living fossil due to its 200 million year’s history under morphological stasis. Its resilience is partly attributed to its unique set of specialized metabolites, in particular, ginkgolides and bilobalide, which are chemically complex terpene trilactones. Here, we use a gene cluster-guided mining approach in combination with co-expression analysis to reveal the primary steps in ginkgolide biosynthesis. We show that five multifunctional cytochrome P450s with atypical catalytic activities generate the tert-butyl group and one of the lactone rings, characteristic of all G. biloba trilactone terpenoids. The reactions include scarless C–C bond cleavage as well as carbon skeleton rearrangement (NIH shift) occurring on a previously unsuspected intermediate. The cytochrome P450s belong to CYP families that diversifies in pre-seed plants and gymnosperms, but are not preserved in angiosperms. Our work uncovers the early ginkgolide pathway and offers a glance into the biosynthesis of terpenoids of the Mesozoic Era.

Ramularia collo-cygni is the causal agent of Ramularia leaf spot disease (RLS) on barley and became, during the recent decades, an increasing threat for farmers across the world. Here, we analyze morphological, transcriptional, and metabolic responses of two barley cultivars having contrasting tolerance to RLS, when infected by an aggressive or mild R. collo-cygni isolate. We found that fungal biomass in leaves of the two cultivars does not correlate with their tolerance to RLS, and both cultivars displayed cell wall reinforcement at the point of contact with the fungal hyphae. Comparative transcriptome analysis identified that the largest transcriptional differences between cultivars are at the early stages of fungal colonization with differential expression of kinases, calmodulins, and defense proteins. Weighted gene co-expression network analysis identified modules of co-expressed genes, and hub genes important for cultivar responses to the two R. collo-cygni isolates. Metabolite analyses of the same leaves identified defense compounds such as p-CHDA and serotonin, correlating with responses observed at transcriptome and morphological level. Together these all-round responses of barley to R. collo-cygni provide molecular tools for further development of genetic and physiological markers that may be tested for improving tolerance of barley to this fungal pathogen.

Significance The hypocretin (Hcrt, also known as orexin) neuropeptides regulate sleep and wake stability, and disturbances of Hcrt can lead to sleep disorders. MicroRNAs (miRNAs) are short noncoding RNAs that fine-tune protein expression levels, and miRNA-based therapeutics are emerging. We report a functional interaction between miRNA (miR-137) and Hcrt . We demonstrate that intracellular miR-137 levels in Hcrt neurons regulate Hcrt expression with downstream effects on wakefulness. Specifically, lowering of miR-137 levels increased wakefulness in mice. We further show that the miR-137:Hcrt interaction is conserved across mice and humans, that miR-137 also regulates sleep–wake balance in zebrafish, and that the MIR137 locus is genetically associated with sleep duration in humans. Together, our findings reveal an evolutionarily conserved sleep–wake regulatory role of miR-137.

35The multicellular embryo, and ultimately the entire organism, is a derivative of the fertilized 36 egg cell. Unlike in animals, transcription factor networks orchestrating faithful egg 37 development are still largely unknown in plants. We have identified that egg cell 38 differentiation in Arabidopsis require interplay between evolutionarily conserved onco-protein 39 homologs RETINOBLASTOMA-RELATED (RBR) and redundant MYB proteins 40 MYB64/MYB119. RBR physically interacts with the MYBs; and with plant-specific 41 transcription factors belonging to the RWP-RK-domain (RKD) family and LEAFY 42 COTYLEDON1 (LEC1), which participate in development of egg cells and inherent stress 43 response. RBR binds to most of these egg cell-expressed loci at the DNA level, partially 44 overlapping with sites of histone methylation H3K27me3. Since deregulation of RKDs 45 phenocopies mutants of RBR and the MYBs in terms of cell proliferation in the egg cell 46 spatial domain, all the corresponding proteins are likely required to restrict parthenogenetic 47 cell divisions of the egg cells. Cross-talk among these transcription factors, and direct 48 regulation by RBR, govern egg cell development and expression of egg-to-zygotic polarity 49 factors of the WUSCHEL RELATED HOMEOBOX family. Together, a network of RBR-50 centric transcription factors underlies egg cell development and stress response, possibly, in 51 combination with several other predicted nodes. 52 53 54 55 Key words 56 egg cell | transcription factor | RETINOBLASTOMA RELATED | MYB | RKD | stress | 57 parthenogenesis 58 59 4 Author summary 60 61The RETINOBLASTOMA protein is one of the core components of the Eukaryotic 62 cell cycle, and corresponding evolutionary homologs have been implicated not only 63 to repress cell division but also to control differentiation and development. How 64 RETINOBLASTOMA RELATED (RBR) associate with other higher order regulators 65 to control faithful egg cell development in sexual plants is pivotal for manipulation of 66 successful reproduction in general, and engineering of parthenogenesis when 67 asexual or apomictic seed progeny are desirable over sexual plants. Using a suite of 68 molecular methods, we show that a RBR-associated transcription factor network 69 operates to specify egg cells in Arabidopsis. Complex cross-regulation within these 70 transcription factors seems to be necessary for successful maternal egg cell to 71 zygotic transition and reproductive stress response. Detailed genetic analysis 72 implicate that RBR and its interactive partners belonging to MYB and RWP-RK 73 transcription factor families are possibly required to prevent parthenogenesis of the 74 sexual egg cells. Novel RBR networks and stress nodes explained in this study 75 might help to improve our understanding of sexual and asexual reproduction. 76 77 78 79 80 Proper differentiation of the egg cells is pivotal for sexual reproduction as well as 81 parthenogenesis. In flowering plants, the egg cells are terminally differentiated within 82 the miniature ...