Arabidopsis TITAN LIKE is required for U12-type intron splicing, especially for AT-AC subtypes

Abstract

Many eukaryotes possess two types of spliceosomes: the U2-dependent and U12-dependent spliceosomes. The U2-dependent spliceosome processes more than 99% of all introns, whereas the U12-dependent spliceosome acts on only about 0.3% of introns, one-third of which start with AT and end with AC, with the remainder having GT-AG termini. How the U12-dependent spliceosome splices two types of introns with different terminal sequences remains poorly understood. Human CENATAC is a subunit of the U12-dependent spliceosome that is particularly required for the splicing of the AT-AC subtype. The Arabidopsis genome contains a single homolog, TITAN LIKE (TTL), but its function in splicing remains unknown. Here, we generated TTL mutants and isolated two viable alleles, of which we analyzed one, designated ttl-142, to investigate TTL function in splicing. ttl-142 carries a 42-nucleotide deletion that removes 14 amino acid residues from the predicted protein, and homozygous mutants exhibit morphological abnormalities. Most U12-dependent introns were less efficiently spliced in ttl-142 than in the wild-type, with the splicing of AT-AC introns particularly suppressed. Splicing suppression in ttl-142 was more extensive than that in a DROL1 mutant, which carries a mutation in a gene specifically required for AT-AC intron splicing. Conversely, fewer genes showed altered expression levels in ttl-142 than in drol1, and most differentially expressed genes differed between the two mutants. These results suggest that the phenotypes of ttl-142 and drol1 mutants may reflect the impairment of distinct spliceosomal functions.