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bioRxiv · 10.1101/2025.09.26.678732

Cleavage of MEP-1 by DPF-3 Reveals Novel Substrate Specificity and Its Impact on Reproductive Fitness

Aygun, I.·Amanullah, A.·Seebacher, J.·Hess, D.·Soneson, C.·Grosshans, H.·Gudipati, R. K.
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Abstract

Proteases are enzymes that catalyse the hydrolysis of peptide bonds in proteins for their functional, modification or degradation. Members of the Dipeptidyl Peptidase IV (DPPIV) family are exopeptidases that cleave dipeptides off the N-termini of their substrate peptides, typically after proline or alanine. Recently, we showed that human DPP4 and Caenorhabditis elegans DPF-3 have a larger target repertoire in vitro, permitting cleavage after additional amino acids. Here, we use TAILS (Terminal Amine Isotopic Labelling of Substrates) to identify DPF-3 targets in vivo and observe cleavage of MEP-1 after threonine, confirming a broader substrate specificity of DPF-3 also in vivo. Demonstrating physiological relevance, we show that rendering MEP-1 resistant to cleavage disrupts its stability, leading to developmental abnormalities such as defective gonadal migration and reproductive issues. Collectively, our findings highlight a previously unappreciated complexity in the substrate specificity of DPPIV family proteases and suggest that their physiological roles may extend beyond what is currently known. IMPORTANTO_LIManuscripts submitted to Review Commons are peer reviewed in a journal-agnostic way. C_LIO_LIUpon transfer of the peer reviewed preprint to a journal, the referee reports will be available in full to the handling editor. C_LIO_LIThe identity of the referees will NOT be communicated to the authors unless the reviewers choose to sign their report. C_LIO_LIThe identity of the referee will be confidentially disclosed to any affiliate journals to which the manuscript is transferred. C_LI GUIDELINESO_LIFor reviewers: https://www.reviewcommons.org/reviewers C_LIO_LIFor authors: https://www.reviewcommons.org/authors C_LI CONTACTThe Review Commons office can be contacted directly at: office@reviewcommons.org

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Aygun, I., Amanullah, A., Seebacher, J., Hess, D., Soneson, C., Grosshans, H., Gudipati, R. K.. 2025-09-28. Cleavage of MEP-1 by DPF-3 Reveals Novel Substrate Specificity and Its Impact on Reproductive Fitness. https://doi.org/10.1101/2025.09.26.678732

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