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Abstract Inhibitory Smads (I-Smads) regulate TGF-β/BMP signaling through multiple distinct mechanisms, but whether different tissues preferentially employ specific mechanisms remains unknown. To address this question, we performed structure-function analyses of the Drosophila I-Smad Dad and its vertebrate orthologs Smad6 and Smad7 in neural and wing tissues, measuring in vivo outputs of BMP signaling. We identified a critical 24-amino acid putative DNA-binding domain (DNABD) within the MH1 domain of the Drosophila I-Smad, Dad, that is essential for inhibitory function in wing tissue but unessential in neural tissue. Structural analyses revealed that ΔDNABD disrupts a beta hairpin structure homologous to R-Smad DNA-binding regions. We also found that Dad requires an intact MH1 domain to disrupt wing development, whereas either MH1 or MH2 can independently disrupt BMP signaling in motor neurons. These findings support a model where Dad functions through MH1-mediated transcriptional regulation in wing primordium, but through multiple mechanisms in neurons. Comparative analysis revealed that vertebrate I-Smad orthologs also show tissue-specific activity patterns, with structural predictions suggesting that Smad6 retains ancestral DNA-binding capacity while Smad7 has evolved enhanced MH2-mediated functions. These results reveal context-dependent mechanisms of I-Smads that further the understanding of TGF-β/BMP pathway regulation.
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ABSTRACT Phenotypic variation is common across life history and among populations occupying different environments, yet the molecular mechanisms underlying these axes of divergence remain poorly understood. Much work has focused on gene expression as a link between genetic variation, environmental variation, and phenotypes, but post‐transcriptional processes such as alternative splicing—which affect how transcripts are assembled rather than how much of a transcript is produced—are increasingly recognized as additional modulators of plasticity and adaptation. Here, we examined gene expression and alternative splicing together in the wood frog ( Rana sylvatica ), an amphibian with a complex life cycle whose populations differ across replicated gradients of road adjacency and associated pollution. We found extensive transcriptomic differences between hatchlings and adults, with thousands of genes differentially expressed or spliced. Individuals clustered strongly by population for both expression and splicing. Differences at the habitat level were less extensive, but revealed two differentially expressed genes ( HSP70 and Gpsm2 ) and one differentially spliced gene ( Cd82 ) that consistently distinguished roadside and woodland populations. Overall, genetic differentiation between populations was low, suggesting that phenotypic and transcriptomic differences likely emerge in the presence of gene flow and reflect plastic responses. Together, these results highlight transcriptomic plasticity as an important mechanism shaping variation across both development and population differentiation.
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- Biology (2)
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- Journal Article (2)
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- English (2)