Dataset Information

Proteases in Quercus ilex seeds

ABSTRACT: Proteases and protease inhibitors have been analyzed in the non-orthodox Quercus ilex forest tree by using in silico and wet approaches. The in silico analysis showed that the species-specific transcriptome database included 2240 and 97 transcripts annotated as, respectively, proteases and protease inhibitors, with members of the different families according to the MEROPS protease database (http://www.merops.ac.uk) classification, mostly of serine- and metallo-type. Data for Q. ilex are compared with those previously published for other Quercus species, namely, Q. suber, Q. lobata, and Q. robur. A second approach was performed to assess the evolution of proteases and protease inhibitors along with seed germination in cotyledon and embryo tissues by determining changes at proteomics and activity levels, the last by mean of in vitro and in gel assays. Shotgun (LC-MSMS) analysis of acorn embryos and cotyledons of three developmental stages (NV, non-viable; T1, mature; and T3 germinated) allowed the identification of a total of 177 proteases and 12 protease inhibitors. In vitro protease activity was determined by using azocasein as substrate; the total activity was higher in cotyledons than embryos, showing the latter significant differences throughout germination. Gel protease activity assays showed common and differential bands between tissues and developmental stages. Shotgun analysis of 10 differential activity bands supported the results obtained from the whole proteome analysis, with some proteins highly represented in the same tissues and stages by both strategies. In overall, the combination of shotgun proteomics and protease activity approaches allowed the identification of tissue-specific (such as cysteine protease inhibitors), and stage-specific proteins (such as those related to storage proteins mobilization), with others being present in all the analyzed organs. Some of the proteases and protease inhibitors differentially identified between tissues and stages in this study could be used as indicators of seed quality and viability.

INSTRUMENT(S): LTQ Orbitrap

ORGANISM(S): Quercus Ilex (holly Oak)

TISSUE(S): Embryo, Cotyledon

SUBMITTER: Maria Angeles Castillejo

LAB HEAD: María Angeles Castillejo Sanchez

PROVIDER: PXD032845 | Pride | 2022-08-11

REPOSITORIES: Pride

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Identification of Proteases and Protease Inhibitors in Seeds of the Recalcitrant Forest Tree Species Quercus ilex.

Escandón Monica M Bigatton Ezequiel D ED Guerrero-Sánchez Victor M VM Hernández-Lao Tamara T Rey Maria-Dolores MD Jorrín-Novo Jesus V JV Castillejo Maria Angeles MA

Frontiers in plant science 20220627

Proteases and protease inhibitors have been identified in the recalcitrant species Quercus ilex using in silico and wet methods, with focus on those present in seeds during germination. In silico analyses showed that the Q. ilex transcriptome database contained 2,240 and 97 transcripts annotated as proteases and protease inhibitors, respectively. They belonged to the different families according to MEROPS, being the serine and metallo ones the most represented. The da ...[more]

PMID: 35832232

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Project description:Proteases constitute the largest enzyme gene family in vertebrates with intracellular and secreted proteases having critical roles in cellular and organ physiology. Intestinal tract contains diverse set of proteases mediating digestion, microbial responses, epithelial and immune signaling. Transit of chyme through the intestinal tract results in significant suppression of proteases. Although endogenous protease inhibitors have been identified, the broader mechanisms underlying protease regulation in the intestinal tract remains unclear. The objective of this study was to determine microbial regulation of proteolytic activity in intestinal tract using phenotype of post-infection irritable bowel syndrome, a condition characterized by high fecal proteolytic activity. Proteases of host pancreatic origin (chymotrypsin like pancreatic elastase 2A, 3B and trypsin 2) drove proteolytic activity. Of the 14 differentially abundant taxa, high proteolytic activity state was characterized by complete absence of the commensal Alistipes putredinis. Germ free mice had very high proteolytic activity (10-fold of specific-pathogen free mice) which dropped significantly upon humanization with microbiota from healthy volunteers. In contrast, high proteolytic activity microbiota failed to inhibit it, a defect that corrected with fecal microbiota transplant as well as addition of A. putredinis. These mice also had increased intestinal permeability similar to that seen in patients. Microbiota β-glucuronidases mediate bilirubin deconjugation and unconjugated bilirubin is an inhibitor of serine proteases. We found that high proteolytic activity patients had lower urobilinogen levels, a product of bilirubin deconjugation. Mice colonized with β-glucuronidase overexpressing E. coli demonstrated significant inhibition of proteolytic activity and treatment with β-glucuronidase inhibitors increased it. The findings establish that specific commensal microbiota mediates effective inhibition of host pancreatic proteases and maintains intestinal barrier function through the production of β-glucuronidases. This suggests an important homeostatic role for commensal intestinal microbiota.

			Action	DRS
	Band1.mgf	Mgf
	Band1.msf	Msf
	Band1.raw	Raw
	Band10.mgf	Mgf
	Band10.msf	Msf

Dataset Information

Proteases in Quercus ilex seeds

Dataset's files

Publications

Identification of Proteases and Protease Inhibitors in Seeds of the Recalcitrant Forest Tree Species <i>Quercus ilex</i>.

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