Rationale:: Mutations in the intercalated disc proteins, such as plakophilin 2 (PKP2), cause arrhythmogenic cardiomyopathy (AC). AC is characterized by the replacement of cardiac myocytes by fibro-adipocytes, cardiac dysfunction, arrhythmias, and sudden death. Objective:: To delineate the molecular pathogenesis of AC. Methods and Results:: Localization and levels of selected intercalated disc proteins, including signaling molecules, were markedly reduced in human hearts with AC. Altered protein constituents of intercalated discs were associated with activation of the upstream Hippo molecules in the human hearts, in Nkx2.5-Cre:Dsp ^W/F and Myh6:Jup mouse models of AC, and in the PKP2 knockdown HL-1 myocytes (HL-1 ^PKP2:shRNA ). Level of active protein kinase C-α isoform, which requires PKP2 for activity, was reduced. In contrast, neurofibromin 2 (or Merlin), a molecule upstream of the Hippo pathway and that is inactivated by protein kinase C-α isoform, was activated. Consequently, the downstream Hippo molecules mammalian STE20-like protein kinases 1/2 (MST1/2), large tumor suppressor kinases 1/2 (LATS1/2), and Yes-associated protein (YAP) (the latter is the effector of the pathway) were phosphorylated. Coimmunoprecipitation detected binding of phosphorylated YAP, phosphorylated β-catenin, and junction protein plakoglobin (the latter translocated from the junction). RNA sequencing, transcript quantitative polymerase chain reaction, and reporter assays showed suppressed activity of SV40 transcriptional enhancer factor domain (TEAD) and transcription factor 7-like 2 (TCF7L2), which are transcription factors of the Hippo and the canonical Wnt signaling, respectively. In contrast, adipogenesis was enhanced. Simultaneous knockdown of Lats1/2 , molecules upstream to YAP, rescued inactivation of YAP and β-catenin and adipogenesis in the HL-1 ^PKP2:shRNA myocytes. Conclusions:: Molecular remodeling of the intercalated discs leads to pathogenic activation of the Hippo pathway, suppression of the canonical Wnt signaling, and enhanced adipogenesis in AC. The findings offer novel mechanisms for the pathogenesis of AC.

Phenylpropanoids are major secondary metabolites in eggplant (Solanum melongena) fruits. Chlorogenic acid (CGA) accounts for 70 to 90% of total phenolics in flesh tissues, while anthocyanins are mainly present in the fruit skin. As a contribution to the understanding of the peculiar accumulation of these health-promoting metabolites in eggplant, we report on metabolite abundance, regulation of CGA and anthocyanin biosynthesis, and characterization of candidate CGA biosynthetic genes in S. melongena. Higher contents of CGA, Delphinidin 3-rutinoside and rutin were found in eggplant fruits compared to other tissues, associated to an elevated transcript abundance of structural genes such as PAL, HQT, DFR and ANS, suggesting that active in situ biosynthesis contributes to anthocyanin and CGA accumulation in fruit tissues. Putative orthologs of the two CGA biosynthetic genes PAL and HQT, as well as a variant of a MYB1 transcription factor showing identity with group 6 MYBs, were isolated from an Occidental S. melongena traditional variety and demonstrated to differ from published sequences from Asiatic varieties. In silico analysis of the isolated SmPAL1, SmHQT1, SmANS, and SmMyb1 promoters revealed the presence of several Myb regulatory elements for the biosynthetic genes and unique elements for the TF, suggesting its involvement in other physiological roles beside phenylpropanoid biosynthesis regulation. Transient overexpression in Nicotiana benthamiana leaves of SmMyb1 and of a C-terminal SmMyb1 truncated form (SmMyb1Δ9) resulted in anthocyanin accumulation only of SmMyb1 agro-infiltrated leaves. A yeast two-hybrid assay confirmed the interaction of both SmMyb1 and SmMyb1Δ9 with an anthocyanin-related potato bHLH1 TF. Interestingly, a doubled amount of CGA was detected in both SmMyb1 and SmMyb1Δ9 agro-infiltrated leaves, thus suggesting that the N-terminal region of SmMyb1 is sufficient to activate its synthesis. These data suggest that a deletion of the C-terminal region of SmMyb1 does not limit its capability to regulate CGA accumulation, but impairs anthocyanin biosynthesis. To our knowledge, this is the first study reporting a functional elucidation of the role of the C-term conserved domain in MYB activator proteins

Rationale:: Mutations in desmosome proteins cause arrhythmogenic cardiomyopathy (AC), a disease characterized by excess myocardial fibroadipocytes. Cellular origin(s) of fibroadipocytes in AC is unknown. Objective:: To identify the cellular origin of adipocytes in AC. Methods and Results:: Human and mouse cardiac cells were depleted from myocytes and flow sorted to isolate cells expressing platelet-derived growth factor receptor-α and exclude those expressing other lineage and fibroblast markers (CD32, CD11B, CD45, Lys76, Ly ^−6c and Ly ^6c , thymocyte differentiation antigen 1, and discoidin domain receptor 2). The PDGFRA ^pos :Lin ^neg :THY1 ^neg :DDR2 ^neg cells were bipotential as the majority expressed collagen 1 α-1, a fibroblast marker, and a subset CCAAT/enhancer-binding protein α, a major adipogenic transcription factor, and therefore, they were referred to as fibroadipocyte progenitors (FAPs). FAPs expressed desmosome proteins, including desmoplakin, predominantly in the adipogenic but not fibrogenic subsets. Conditional heterozygous deletion of Dsp in mice using Pdgfra-Cre deleter led to increased fibroadipogenesis in the heart and mild cardiac dysfunction. Genetic fate mapping tagged 41.4±4.1% of the cardiac adipocytes in the Pdgfra-Cre : Eyfp:Dsp ^{W /F} mice, indicating an origin from FAPs. FAPs isolated from the Pdgfra-Cre : Eyfp:Dsp ^{W /F} mouse hearts showed enhanced differentiation to adipocytes. Mechanistically, deletion of Dsp was associated with suppressed canonical Wnt signaling and enhanced adipogenesis. In contrast, activation of the canonical Wnt signaling rescued adipogenesis in a dose-dependent manner. Conclusions:: A subset of cardiac FAPs, identified by the PDGFRA ^pos :Lin ^neg :THY1 ^neg :DDR2 ^neg signature, expresses desmosome proteins and differentiates to adipocytes in AC through a Wnt-dependent mechanism. The findings expand the cellular spectrum of AC, commonly recognized as a disease of cardiac myocytes, to include nonmyocyte cells in the heart.

The role of calcineurin protein phosphatase 2B (PP2B) in the pathogenesis of human hypertrophic cardiomyopathy (HCM) remains unsettled. We determined potential involvement of calcineurin in the pathogenesis of HCM caused by mutations in myozenin 2 (MYOZ2), an inhibitor of calcineurin. We generated multiple lines of transgenic mice expressing either Flag-tagged wild-type (WT) (MYOZ2^WT) or mutant MYOZ2^S48P and MYOZ2^I246M, identified in families with HCM, in the heart. To mimic the human genotype, we generated bigenic mice expressing WT and mutant MYOZ2 in the background of hemizygous endogenous MYOZ2 (Myoz2^+/−). Transgene proteins constituted 15–48% of the total MYOZ2 protein in the heart. Mutant MYOZ2 mice showed molecular, cellular, and gross cardiac hypertrophy, preserved systolic function, and interstitial fibrosis. Immunofluorescence staining showed co-localization of WT and mutant MYOZ2 proteins with α-actinin at the Z disks. Electron microscopy showed disrupted and mal-aligned Z disks in the mutant mice. Cardiac calcineurin activity, determined by quantifying Rcan1.4 mRNA and protein levels, luciferase activity in triple transgenic Myoz2^+/−:NFATc-Luc:MYOZ2^I246M and Myoz2^+/−:NFATc-Luc:MYOZ2^WT mice, and NFATc transcriptional activity assay, was unchanged in the mutant transgenic mice. However, levels of phospho-ERK1/2 and JNK54/46 were altered in the transgenic mice. Likewise, lentiviral-mediated expression of the MYOZ2^I246M did not affect RCAN1.4 and calcineurin (PPP3CB) protein levels. Thus, the cardiac phenotype in HCM caused by MYOZ2 mutations might be independent of calcineurin activity in the heart. Z disk abnormalities might provide the stimulus for the induction of cardiac hypertrophy caused by MYOZ2 mutations.

Summary: Modulation of growth in response to environmental cues is a fundamental aspect of plant adaptation to abiotic stresses. TIP41 (TAP42 INTERACTING PROTEIN OF 41 kDa) is the Arabidopsis thaliana orthologue of proteins isolated in mammals and yeast that participate in the Target‐of‐Rapamycin (TOR) pathway, which modifies cell growth in response to nutrient status and environmental conditions. Here, we characterized the function of TIP41 in Arabidopsis. Expression analyses showed that TIP41 is constitutively expressed in vascular tissues, and is induced following long‐term exposure to NaCl, polyethylene glycol and abscisic acid (ABA), suggesting a role of TIP41 in adaptation to abiotic stress. Visualization of a fusion protein with yellow fluorescent protein indicated that TIP41 is localized in the cytoplasm and the nucleus. Abolished expression of TIP41 results in smaller plants with a lower number of rosette leaves and lateral roots, and an increased sensitivity to treatments with chemical TOR inhibitors, indicating that TOR signalling is affected in these mutants. In addition, tip41 mutants are hypersensitive to ABA at germination and seedling stage, whereas over‐expressing plants show higher tolerance. Several TOR‐ and ABA‐responsive genes are differentially expressed in tip41, including iron homeostasis, senescence and ethylene‐associated genes. In yeast and mammals, TIP41 provides a link between the TOR pathway and the protein phosphatase 2A (PP2A), which in plants participates in several ABA‐mediated mechanisms. Here, we showed an interaction of TIP41 with the catalytic subunit of PP2A. Taken together, these results offer important insights into the function of Arabidopsis TIP41 in the modulation of plant growth and ABA responses.

Arabidopsis DNA-DAMAGE REPAIR/TOLERATION PROTEIN111/SPLICING FACTOR FOR PHYTOCHROME SIGNALING regulates pre-mRNA splicing in seeds, affecting ABA-mediated respo