A bona fide La protein is required for embryogenesis in Arabidopsis thaliana

Searches in the Arabidopsis thaliana genome using the La motif as query revealed the presence of eight La or La-like proteins. Using structural and phylogenetic criteria, we identified two putative genuine La proteins (At32 and At79) and showed that both are expressed throughout plant development but at different levels and under different regulatory conditions. At32, but not At79, restores Saccharomyces cerevisiae La nuclear functions in non-coding RNAs biogenesis and is able to bind to plant 3′-UUU-OH RNAs. We conclude that these La nuclear functions are conserved in Arabidopsis and supported by At32, which we renamed as AtLa1. Consistently, AtLa1 is predominantly localized to the plant nucleoplasm and was also detected in the nucleolar cavity. The inactivation of AtLa1 in Arabidopsis leads to an embryonic-lethal phenotype with deficient embryos arrested at early globular stage of development. In addition, mutant embryonic cells display a nucleolar hypertrophy suggesting that AtLa1 is required for normal ribosome biogenesis. The identification of two distantly related proteins with all structural characteristics of genuine La proteins suggests that these factors evolved to a certain level of specialization in plants. This unprecedented situation provides a unique opportunity to dissect the very different aspects of this crucial cellular activity

Caractérisation de la fonction "La" chez Arabidopsis thaliana et identification d'une structure conservée pour les ARN non-codants de type SINE

SINEs are mobile DNA elements found in almost all eukaryotes. Study of SINEs first focused on their retroposition mechanism, mutagenic effect and general impact on the structure and evolution of genomes. However SINE RNAs have recently been proposed to act as cellular riboregulators. Studying cis and trans elements taking part in the metabolism of SINE RNA in the model plant Arabidopsis thaliana, we aim to better understand SINE element biology. First, we experimentally defined the secondary structure of two tRNA-related SINE RNA: SB1 from Brassica napus and SB2 from Arabidopsis. Although unrelated at the primary sequence level, we found that these RNAs present similar secondary structures. Following this observation, an in silico analysis including tRNA-related SINE RNAs from various eukaryotes has been performed by FJ Sun, G Caetano-Anollés and JM Deragon. This study underlines the existence of common evolutionary trends for SINE RNA secondary structure that could be linked with the SINE RNA riboregulators function. Searching for trans factors involved in SINE RNA metabolism, we have chosen to characterise the La protein, an ubiquitous RNA-binding protein involved in the metabolism of various RNAs, from non-coding RNA to cellular or viral mRNA. Unlike other eukaryotes, which have only one La protein, we identified in Arabidopsis two proteins with the phylogenetic and structural characteristics of genuine La protein: At32 and At79. We showed that At32 (renamed AtLa1) is able to fulfil La nuclear functions in non-coding RNA maturation, including SINE RNA. We also demonstrate that loss of AtLa1 function leads to embryonic lethality. Although AtLa1 and At79 have the same nuclear localisation, loss of At79 function did not affect viability. AtLa1 and At79 have differing levels and profiles of expression. Furthermore, the AtLa1 and At79 proteins apparently bind distinct sets of RNA. We thus propose that Arabidopsis possess two functional homologues of the La protein, which have partially specialised to fulfil different aspects of the La function.L'étude des éléments cis et trans intervenant dans le métabolisme des ARN SINE a pour but de mieux appréhender la biologie des éléments mobiles SINE. Nous avons d'une part déterminé la structure secondaire des ARN des SINE de plante SB1 et SB2. Ces deux ARN qui n'ont pas d'homologie de séquence adoptent des structures secondaires similaires. Suite à cette observation, une étude menée par F.J. Sun et al. a mis en évidence un schéma évolutif commun des ARN SINE dérivés d'ARNt. Dans le cadre de la recherche de facteurs trans, nous avons entrepris la caractérisation de la protéine La, un facteur de liaison à l'ARN. Nous avons identifié chez Arabidopsis deux protéines présentant toutes les caractéristiques de la protéine La : At32 et At79. Nous avons montré que seule At32 assure les fonctions nucléaires de la protéine La. At32 et At79 ont des profils d'expression différents et semblent lier des ARN distincts. Nous proposons donc qu'il existe deux homologues de la protéine La Chez Arabidopsi