OVATE family protein 20 regulates fruit shoulder width by modulating auxin distribution in pepper

Fruit shape is a key determinant of pepper (Capsicum annuum L.) fruit commercial value, and the shoulder is a crucial feature of the fruit's proximal end. Despite the fruit shoulder being an important parameter in morphological variation, the genes and regulatory mechanisms underlying fruit shoulder morphology remain elusive. Here, we identified a fruit shoulder (fs) mutant that primarily exhibits a shoulderless fruit phenotype, with an ∼40% reduction in fruit weight. Our data demonstrated that OVATE family protein 20 (CaOFP20) controls this phenotype. Silencing CaOFP20 in various fruit types consistently led to decreased fruit shoulder width, while fruit length was unaffected. CaOFP20 was mainly expressed in the pericarp, seeds, and placenta, where it regulates organ size by modulating cell division. Loss of CaOFP20 disrupted the expression of auxin pathway genes and the spatial distribution of auxin in fruits. Additionally, we demonstrated that CaOFP20 physically interacts with HECATE1 (CaHEC1), and silencing CaHEC1 in fs resulted in a wider fruit shoulder. Moreover, CaOFP20 and CaHEC1 play distinct yet complementary roles in regulating the expression of auxin pathway genes involved in fruit shoulder development, such as PIN-FORMED1 (CaPIN1), CaPIN3, and Indole-3-Acetic Acid Inducible (CaIAA9), thus finely controlling fruit shoulder development. Taken together, our findings establish CaOFP20 as a central regulator of fruit shoulder development and suggest its potential utility in breeding programs to enhance pepper fruit quality.