Peptidomics Methods Applied to the Study of Flower Development

Understanding the global and dynamic nature of plant developmental processes requires not only the study of the transcriptome, but also of the proteome, including its largely uncharacterized peptidome fraction. Recent advances in proteomics and high-throughput analyses of translating RNAs (ribosome profiling) have begun to address this issue, evidencing the existence of novel, uncharacterized, and possibly functional peptides. To validate the accumulation in tissues of sORF-encoded polypeptides (SEPs), the basic setup of proteomic analyses (i.e., LC-MS/MS) can be followed. However, the detection of peptides that are small (up to ~100 aa, 6–7 kDa) and novel (i.e., not annotated in reference databases) presents specific challenges that need to be addressed both experimentally and with computational biology resources. Several methods have been developed in recent years to isolate and identify peptides from plant tissues. In this chapter, we outline two different peptide extraction protocols and the subsequent peptide identification by mass spectrometry using the database search or the de novo identification methods.

Our work on peptidomics was supported by grant BFU2014-58289-P (funded by MICIN/AEI/ 10.13039/501100011033 and by “ERDF A way of making Europe”) and by grant 2017SGR718 (from the Agencia de Gestió d’Ajuts Universitaris I de Recerca) to JLR, and by institutional grant SEV-2015-0533 (funded by MCIN/AEI/10.13039/501100011033) and by the CERCA Programme/Generalitat de Catalunya. R.A. is supported by fellowship PRE2018-084278 funded by MCIN/AEI/10.13039/501100011033 and by “ESF Investing in your future.” The CRG/UPF Proteomics Unit is part of the Spanish Infrastructure for Omics Technologies (ICTS OmicsTech). We also acknowledge “Secretaria d’Universitats i Recerca del Departament d’Economia i Coneixement de la Generalitat de Catalunya” (2017SGR595) and support of the Spanish Ministry of Science and Innovation to the EMBL partnership, the Centro de Excelencia Severo Ochoa, and the CERCA Programme/Generalitat de Catalunya.

Peer reviewed