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The inability to maintain a strictly regulated endo(lyso)somal acidic pH through the proton-pumping action of the vacuolar-ATPases (v-ATPases) has been associated with various human diseases including heritable connective tissue disorders. Autosomal recessive (AR) cutis laxa (CL) type 2C syndrome is associated with genetic defects in the ATP6V1E1 gene and is characterized by skin wrinkles or loose redundant skin folds with pleiotropic systemic manifestations. The underlying pathological mechanisms leading to the clinical presentations remain largely unknown. Here, we show that loss of atp6v1e1b in zebrafish leads to early mortality, associated with craniofacial dysmorphisms, vascular anomalies, cardiac dysfunction, N-glycosylation defects, hypotonia, and epidermal structural defects. These features are reminiscent of the phenotypic manifestations in ARCL type 2C patients. Our data demonstrates that loss of atp6v1e1b alters endo(lyso)somal protein levels, and interferes with non-canonical v-ATPase pathways in vivo. In order to gain further insights into the processes affected by loss of atp6v1e1b, we performed an untargeted analysis of the transcriptome, metabolome, and lipidome in early atp6v1e1b-deficient larvae. We report multiple affected pathways including but not limited to oxidative phosphorylation, sphingolipid, fatty acid, and energy metabolism together with profound defects on mitochondrial respiration. Taken together, our results identify complex pathobiological effects due to loss of atp6v1e1b in vivo., Author summary Cutis laxa syndromes are pleiotropic disorders of the connective tissue, characterized by skin redundancy and variable systemic manifestations. Cutis laxa syndromes are caused by pathogenic variants in genes encoding structural and regulatory components of the extracellular matrix or in genes encoding components of cellular trafficking, metabolism, and mitochondrial function. Pathogenic variants in genes coding for vacuolar-ATPases, a multisubunit complex responsible for the acidification of multiple intracellular vesicles, cause type 2 cutis laxa syndromes, a group of cutis laxa subtypes further characterized by neurological, skeletal, and rarely cardiopulmonary manifestations. To investigate the pathomechanisms of vacuolar-ATPase dysfunction, we generated zebrafish models that lack a crucial subunit of the vacuolar-ATPases. The mutant zebrafish models show morphological and functional features reminiscent of the phenotypic manifestations in cutis laxa patients carrying pathogenic variants in ATP6V1E1. In-depth analysis at multiple -omic levels identified biological signatures that indicate impairment of signaling pathways, lipid metabolism, and mitochondrial respiration. We anticipate that these data will contribute to a better understanding of the pathogenesis of cutis laxa syndromes and other disorders involving defective v-ATPase function, which may eventually improve patient treatment and management.

Background Chromatin immunoprecipitation combined with massive parallel sequencing (ChIP-seq) is widely used to study protein-chromatin interactions or chromatin modifications at genome-wide level. Sequence reads that accumulate locally at the genome (peaks) reveal loci of selectively modified chromatin or specific sites of chromatin-binding factors. Computational approaches (peak callers) have been developed to identify the global pattern of these sites, most of which assess the deviation from background by applying distribution statistics. Results We have implemented MeDiChISeq, a regression-based approach, which - by following a learning process - defines a representative binding pattern from the investigated ChIP-seq dataset. Using this model MeDiChISeq identifies significant genome-wide patterns of chromatin-bound factors or chromatin modification. MeDiChISeq has been validated for various publicly available ChIP-seq datasets and extensively compared with other peak callers. Conclusions MeDiChI-Seq has a high resolution when identifying binding events, a high degree of peak-assessment reproducibility in biological replicates, a low level of false calls and a high true discovery rate when evaluated in the context of gold-standard benchmark datasets. Importantly, this approach can be applied not only to ‘sharp’ binding patterns - like those retrieved for transcription factors (TFs) - but also to the broad binding patterns seen for several histone modifications. Notably, we show that at high sequencing depths, MeDiChISeq outperforms other algorithms due to its powerful peak shape recognition capacity which facilitates discerning significant binding events from spurious background enrichment patterns that are enhanced with increased sequencing depths. Electronic supplementary material The online version of this article (doi:10.1186/1471-2164-14-834) contains supplementary material, which is available to authorized users.

Linear amplification of DNA (LinDA) by T7 polymerase is a versatile and robust method for generating sufficient amounts of DNA for genome-wide studies with minute amounts of cells. LinDA can be coupled to a great number of global profiling technologies. Indeed, chromatin immunoprecipitation coupled to massive parallel sequencing (ChIP-seq) has been achieved for transcription factors and epigenetic modification of chromatin histones with 1,000 to 5,000 cells. LinDA largely simplifies reChIP-seq experiments to monitor co-binding at chromatin target sites. The single-tube design of LinDA is ideal for handling ultrasmall amounts of DNA (