Characterization of three alternative splice variants associated with theArabidopsisrhomboid protein geneAt1g74130

Abstract

Rhomboid serine proteases are grouped into three main types -secretases, presenilin-like associated rhomboid-like (PARL) proteases, and "inactive" rhomboid proteins. Although the three rhomboid groups are distinct, the different types are likely to operate within the same cell or compartment, such as observed in the plastids of Arabidopsis. There are four distinct plastid rhomboid genes at play in Arabidopsis plastids, two for active types (At1g25290 and At5g25752) and two for inactive forms (At1g74130 and At1g74140). The number of working plastid rhomboids is further increased by alternative splicing, as reported for At1g25290. To understand how the plastid rhomboid system works, it is necessary to identify all rhomboid forms in play. To this end, this study was designed to examine the alternative splicing activities of At1g74130, one of the two genes encoding proteolytically "inactive" plastid rhomboids. The exon mapping and DNA sequencing results obtained here indicate the presence of three prominent alternative splice variants in the At1g74130 transcript population. The dominant splice variant, L, encodes the full-length protein. The other two splice variants, M and S, produce proteins lacking sections from the carboxyl transmembrane domain region. The splice variants M and S appear to be at levels with functional potential and appear to adjust relative to each other during development and in response to changes in the level of Tic40, a component of the plastid translocon. The splice variant proteins themselves exhibit different characteristics with respect to rhomboid protein-substrate interactions. These differences were observed in bacterial co-expression pull-down assays and in yeast mitochondrial studies. When considered together, the data suggest that the alternative splicing of At1g74130 bears functional significance in Arabidopsis and is likely to be part of a mechanism for diversifying plastid rhomboid function. Résumé :On regroupe les protéases sérines rhomboïdes en trois types principaux : secrétases, protéases d'aspect rhomboïdes associées à des présélinines et protéines rhomboïdes « inactives ». Bien que les trois groupes rhomboïdes soient distincts, les différents types agissent probablement dans la même cellule ou compartiment, tel qu'on l'observe dans les plastes de l'Arabidopsis. Quatre gènes rhomboïdes plastidiques distincts agissent dans les plastes de l'Arabidopsis, deux pour les types actifs (At1g25290 et At5g25752) et deux pour les formes inactives (At1g74130 et At1g74140). Le nombre de rhomboïdes plastidiques fonctionnels augmente encore par épissage alternatif, comme rapporté pour le At1g25290. Afin de comprendre comment le système de rhomboïdes plastidiques fonctionne, on doit identifier toutes les formes rhomboïdes en jeu. À cette fin, les auteurs ont conçu cette étude de manière à examiner les activités d'épissage alternatives du At1g74130, un des deux gènes codant pour les rhomboïdes plastidiques protéolytiquement « inactifs ». Les résultats obtenus de la cartographie des exons et ...

Inactive rhomboid proteins: New mechanisms with implications in health and disease

“…Moreover, automated gene annotation by hidden Markov models, and BLAST searches, identified several more distantly related catalytically inactive rhomboid relatives amongst the entire eukaryotic kingdom including, fungi, plants and red algae [4,6,[8][9][10].…”