Correcting signal biases and detecting regulatory elements in STARR-seq data

High-throughput reporter assays such as self-transcribing active regulatory region sequencing (STARR-seq) have made it possible to measure regulatory element activity across the entire human genome at once. The resulting data, however, present substantial analytical challenges. Here, we identify technical biases that explain most of the variance in STARR-seq data. We then develop a statistical model to correct those biases and to improve detection of regulatory elements. This approach substantially improves precision and recall over current methods, improves detection of both activating and repressive regulatory elements, and controls for false discoveries despite strong local correlations in signal.

Gene regulation is of foundational importance to nearly all biological processes, and variation in gene regulatory activity plays a major role in human disease risk (Lee and Young 2013; Parker et al. 2013; Finucane et al. 2015). A major step toward measuring regulatory activity across the human genome has been the development of high-throughput reporter assays such as STARR-seq (Arnold et al. 2013) that allow regulatory element activity to be quantified with high-throughput sequencing rather than with optical detection of a fluorescent or luminescent signal.High-throughput reporter assays create substantial analytical challenges that are distinct from other sequencing-based genomic assays. There is significant local variation in high-throughput reporter assay signal. We show here that, across data from several laboratories, most of that variation can be explained by features of the underlying genomic sequence and experimental procedures rather than by regulatory element activity. For example, nucleotide composition can alter PCR efficiency leading to under- and overrepresentation of some sequences. Meanwhile, highly repetitive sequences often do not align uniquely to the human reference genome, also biasing signal estimates. Additional analytical challenges include that STARR-seq signals can be both positive and negative, reflecting activation and repression, and the boundaries of regulatory elements are typically unknown and must therefore be estimated from the data. Those challenges together impact signal representations, hinder estimation of regulatory element activity, and cause false positives and false negatives when left unaddressed.Taken together, key requirements of statistical methods to analyze STARR-seq data are the ability to identify and estimate the effect of both activating and repressing regulatory elements while also correcting for underlying sequence biases in high-throughput reporter assays. A statistical model was recently introduced that corrects technical biases and detects regulatory elements in STARR-seq, but the model is limited to detecting only activating regulatory elements (Lee et al. 2020). Considering repression is a crucial gene regulation mechanism (Courey and Jia 2001), overlooking repressive elements may limit understanding of gene regulation with STARR-seq. To overcome that challenge, our correcting reads and analysis of differentially active elements (CRADLE) model takes a two-step approach. First, CRADLE uses a generalized linear regression model to estimate and correct major biases that we have identified in STARR-seq data. Next, CRADLE detects regions with statistically significant regulatory activity from the bias-corrected signals while rigorously controlling FDR. In doing so, CRADLE substantially improves the use of STARR-seq by providing a robust estimation of regulatory activity and improved visualization of raw signals.     

25-Hydroxyvitamin D and Cardiorespiratory Fitness in Prepubertal Overweight and Obese Children

Childhood obesity has become a major global health problem. Vitamin D deficiency and poor cardiorespiratory fitness are highly prevalent in children with overweight or obesity, but little is known about their relationships. In this study, we aimed to analyze the relationship between serum 25-hydroxyvitamin D (25(OH)D) and cardiorespiratory fitness parameters in prepubertal obese and overweight children. A cross-sectional design with a sample of 57 prepubertal children, aged 9–11 years, with overweight or obesity was used. The fasting concentration of 25(OH)D was analyzed with a chemiluminescent microparticle immunoassay. Fat and lean body masses were determined by using DXA. Maximal oxygen uptake (VO_2max) was measured with the maximal treadmill test. A total of 68.4% of the sample had sufficient levels of 25(OH)D. As expected, their cardiorespiratory fitness was poor compared with that of normal-weight children, but 60% of the group exceeded the median obesity-specific reference values. No differences were found between the sexes for relative VO_2max or 25(OH)D levels. Moreover, no correlations were found between 25(OH)D and body composition or cardiorespiratory parameters for sex or vitamin D groups. Vitamin D status seems not to be directly related to body composition or cardiorespiratory fitness in prepubertal overweight or obese children.     

Erratum zu: Digital Divide – Soziale Unterschiede in der Nutzung digitaler Gesundheitsangebote

Bundesgesundheitsblatt - Gesundheitsforschung - Gesundheitsschutz - Ein Erratum zu dieser Publikation wurde veröffentlicht: https://doi.org/10.1007/s00103-019-03081-y     

Creating patient-centered radiology reports to empower patients undergoing prostate magnetic resonance imaging

IntroductionAs we progress to an era when patient autonomy and shared decision-making are highly valued, there is a need to also have effective patient-centered communication tools. Radiology reports are designed for clinicians and can be very technical and difficult for patients to understand. It is important for patients to understand their magnetic resonance imaging (MRI) report in order to make an informed treatment decision with their physician. Therefore, we aimed to create a patient-centered prostate MRI report to give our patients a better understanding of their clinical condition.MethodsA prototype patient-centered radiology report (PACERR) was created by identifying items to include based on opinions sought from a group of patients undergoing prostate MRI and medical experts. Data was collected in semi-structured interviews using a salient belief question. A prototype PACERR was created in collaboration with human factors engineering and design, medical imaging, biomedical informatics, and cancer patient education groups.ResultsFifteen patients and eight experts from urology, radiation oncology, radiology, and nursing participated in this study. Patients were particularly interested to have a report with laymen terms, concise language, contextualization of values, definitions of medical terms, and next course of action. Everyone believed the report should include the risk of MRI findings actually being cancer in the subsequent biopsy.ConclusionsA prostate MRI PACERR has been developed to communicate the most important findings relevant to decision-making in prostate cancer using patient-oriented design principles. The ability of this tool to improve patient knowledge and communication will be explored.     

Design Characteristics for the Tissue Engineering of Cartilaginous Tissues

Tissues like the temporomandibular joint (TMJ) disc and the knee meniscus are often mistakenly viewed as a tantamount to hyaline cartilage, largely due to the absence of a comprehensive understanding of the distinguishing properties of cartilaginous tissues. Because of this confusion, fibrocartilaginous tissue engineering attempts may not be based on suitable experimental designs. Fibrocartilaginous tissues are markedly different than hyaline cartilage; however, the dearth of knowledge related to their cellular and biochemical composition, as well as their biomechanical characteristics, is stunning. Hyaline articular cartilage is exclusively composed of chondrocytes that produce primarily type II collagen, whereas the TMJ disc and the knee meniscus have a mixed cell population of fibroblasts and cells similar to chondrocytes, which predominantly secrete type I collagen. Additionally, fibrocartilaginous tissues have a low glycosaminoglycan content, a low compressive modulus, and a high tensile modulus when compared to hyaline cartilage. Therefore, it is crucial for fibrocartilaginous tissue engineering attempts to be tissue-specific, utilizing the knowledge of the distinct and unique properties of these tissues. At the same time, advances and insights related to the science and engineering aspect of hyaline cartilage regeneration must be carefully considered for the in vitro engineering of fibrocartilaginous tissues.     

Induction of p53 up-regulated modulator of apoptosis messenger RNA by chemotherapeutic treatment of locally advanced breast cancer.

PURPOSE: In biopsies of patients with locally advanced breast cancer, we investigated the in vivo changes of the gene expression pattern induced by chemotherapy to find genes that are potentially responsible for the efficacy of the drug. EXPERIMENTAL DESIGN: Early cellular responses to chemotherapy-induced damage, both in vivo and in vitro, were investigated by analyzing chemotherapy-induced changes in gene expression profiles. Core biopsies were taken from nine patients with locally advanced breast cancer, before and at 6 hours after initiation of doxorubicin-based chemotherapy. Both samples were cohybridized on the same microarray containing 18,000 cDNA spots. RESULTS: The analysis revealed marked differences in gene expression profile between treated and untreated samples. The gene which was most frequently found to be differentially expressed was p53 up-regulated modulator of apoptosis (PUMA). This gene was up-regulated in eight of nine patients with an average factor of 1.80 (range, 1.36-2.73). In vitro MCF-7 breast cancer cells exposed to clinically achievable doxorubicin concentrations for 6 hours revealed marked induction of PUMA mRNA, as well. CONCLUSIONS: This is the first report describing PUMA mRNA to be up-regulated as a response to chemotherapy in patients. Because PUMA is a known member of the family of BH3-only proapoptotic proteins, this finding suggests PUMA's potential importance for the response to anticancer drugs.