Adaptive evolution of <i>BMP4</i> as a potential mechanism for flipper forelimb changes in cetaceans

Vertebrate limbs have undergone profound morphological diversification, enabling adaptations to a broad spectrum of ecological niches. In marine mammals, the evolution of highly specialized flipper-like forelimbs represents a profound structural transformation associated with aquatic habitats. This adaptation has been hypothesized to result, in part, from the inhibition of interphalangeal cell apoptosis during limb development, although the underlying genetic mechanism remains poorly understood. This study investigated the evolutionary dynamics and functional consequences of three key bone morphogenetic protein genes, <i>BMP2</i>, <i>BMP4</i>, and <i>BMP7</i>, which regulate apoptosis in interphalangeal mesenchymal stromal cells during embryonic limb development to ensure proper differentiation of interphalangeal tissues. Comparative genomic analysis revealed significantly accelerated evolution for <i>BMP4</i> and <i>BMP7</i> in the cetacean ancestral lineage, with two positively selected sites (V79I and H247R) involved in cetacean-specific amino acid substitutions located in the TGF-β propeptide functional domain in <i>BMP4</i>. <i>In vitro</i> assays confirmed that cetacean-specific <i>BMP4</i> mutations significantly disrupted normal cell apoptosis and proliferation and altered the transcription and protein expression of downstream apoptosis-related factors, including cytochrome c (Cyt c), BCL2 associated X, and B-cell lymphoma 2, within the BMP signaling pathway. The significant influence of <i>BMP4</i> mutations on apoptotic inhibition highlights a potential role in the development of limb bud mesenchymal tissue and the emergence of the flipper forelimb phenotype in cetaceans.