Palindromi i potencijalne sekundarne strukture u transkriptima izoformi biljnih glutamin-sintetaza

Abstract

Glutamine synthetase (GS, E.C. 6.3.1.2) plays a central role in nitrogen metabolism by assimilating ammonia into glutamine. Plants have one chloroplastic (GS2) and one or more cytosolic (GS1) isoforms. Zea mays and Arabidopsis thaliana are among species with largest GS gene families, having one GS2 and five GS1 genes. Regulation of expression of GS genes, as well as regulation of GS enzymes at the posttranslational level has been widely studied. However, there is limited information on the possibilities of posttranscriptional regulation at the level of GS transcripts. Hereby we show that maize GS mRNAs may adopt stabile secondary structures in the form of hairpins and homo- or heterodimers in the region preceding or including their AUG codon, which may influence translation efficiency and transcript stability. The possibility of forming hairpins is based on the presence of palindromes with gaps, while dimers can form if sequences contain palindromes without gaps. All six maize GS transcripts, as well as three Arabidopsis GS transcripts have one 6-nt palindrome without gap, while one Arabidopsis transcript has two gapless 6-nt palindromes. In addition, maize GS1-3 and Arabidopsis GLN1;2 isoforms contain a perfect 8-nt palindrome with a gap. To show statistical significance of these findings, the number of found palindromes in GS transcripts was expressed as a ratio of found and expected number of palindromes of all sizes, with and without gaps in sequences of a given length. Our bioinformatics analysis suggests that GS transcripts contain structural features that allow regulation at the level of mRNAs, and presents a good starting point for experimental studies of the proposed posttranslational regulation.

Glutamin sintetaza (GS, E.C. 6.3.1.2) ima centralnu ulogu u metabolizmu azota jer asimiluje amonijak u glutamin. Biljke poseduju jednu hloroplastnu (GS2) i jednu ili više citosolnih (GS1) izoformi. Zea mays i Arabidopsis thaliana spadaju u vrste sa najvećim familijama GS gena, jer pored jedne GS2 imaju po pet GS1 izoformi. Regulacija ekspresije GS gena, kao i regulacija enzimske GS aktivnosti na posttranslacionom nivou mnogo je proučavana. Međutim, gotovo da i nema informacija o mogućnosti posttranskripcione regulacije na nivou GS transkripata. Mi smo pokazali da GS mRNK mogu da poprime stabilne sekundarne strukture ukosnice ili homo i heterodimera u regionu koji prethodi ili uključuje AUG kodon, što može uticati na efikasnost translacije i na stabilnost transkripata. Mogućnost formiranja ukosnica zasniva se na prisustvu palindroma sa razmacima, dok se dimeri mogu formirati ukoliko sekvence sadrže palindome bez razmaka. Svih šest transkripata GS kukuruza, kao i tri transkripta A. thaliana imaju po jedan palindrom od 6 nt bez razmaka, dok jedan transkript A. thaliana ima dva ovakva palindroma. Pored toga, GS1-3 kukuruza i GLN1;2 A. thaliana imaju po jedan perfektan palindrom od 8 nt sa razmakom. Kako bi se pokazao statistički značaj ovih nalaza, broj nađenih palindroma u GS transkriptima predstavljen je kao odnos broja nađenih i broja očekivanih palindroma svih dužina sa i bez razmaka u sekvencama datih dužina. Naša bioinformatička analiza sugeriše da GS transkripti poseduju strukturne elemente koji omogućavaju regulaciju na nivou mRNK i predstavlja dobru polaznu osnovu za eksperimentalne studije predložene posttranslacione regulacije.