The causes of most human birth defects are unknown. Structural defects in the heart and digestive tract are frequently found in association with abnormal left-right asymmetries in other organ systems, suggesting that such deformities may result from perturbed laterality, yet the developmental processes that form asymmetries within individual organs remain elusive. The long term goal is to determine the morphogenetic mechanisms that control the development of anatomical left-right asymmetry. The objective of this exploratory R21 application is to gain a comprehensive view of the left-right asymmetric molecular differences within the embryonic heart and gut tubes as they undergo asymmetric "looping", a fundamental organogenesis event that orients the most crucial anatomical asymmetries. To accomplish this objective, the unique embryological features of a novel model amphibian, Lepidobatrachus laevis, will be developed and exploited. Lepidobatrachus has massive embryos that facilitate precise excision of the left and right halves of the early heart an gut during looping, enabling the heretofore infeasible approach of left-right transcriptome profiling during a key phase of asymmetric morphogenesis. The central hypothesis is that identifying transcripts that are differentially expressed between the contralateral halves of looping organs will identify new molecules that control left-right asymmetric morphogenesis. This hypothesis will be tested via two specific aims: 1) Identify transcripts that are differentialy expressed between the left and right sides of the looping heart and gut tubes;and 2) Validate the biological relevance of left- or right-enriched transcripts for asymmetric organ morphogenesis.
Under Aim 1, an RNAseq approach (supported by a draft Lepidobatrachus transcriptome already constructed by the PI) will be used to complete genome-wide expression analyses that will reveal unilaterally-enriched transcripts associated with the formation of key anatomical asymmetries.
Under Aim 2, the proven ability to precisely target exogenous reagents to the left or right side of developing organs in amphibians, and the unprecedented subcellular resolution of developing organ asymmetries provided by the sizeable Lepidobatrachus, will be used to authenticate the in vivo function of select unilaterally-enriched transcripts in asymmetric morphogenesis. The approach is innovative because it takes advantage of the distinctive attributes of a unique non-model organism to understand one of the key unanswered questions in the field of left-right development: what are the mechanism(s) by which developing organs acquire critical left- right asymmetric anatomical features? The proposed research is significant because it is expected to immediately accelerate our understanding of the etiology of some of the most common birth defects by identifying new classes of molecules, and new cellular processes, which shape the fundamental left-right asymmetry of the heart and gut.
Birth defects are the leading cause of infant mortality in the United States, yet most have no known origin. Defining the etiology of these developmental malformations in various organs is of significant clinical importance. The information gained from this study will ultimately be used to detect homologous gene variants in human patient populations, and may identify biomarkers for birth defect risk or therapeutic intervention that could lead to decreased morbidity or mortality in human infants. Importantly, this work will also launch Lepidobatrachus laevis as an exciting new animal model with inimitable experimental advantages for investigating a broad spectrum of developmental biology and disease.