It is generally believed that splicing removes introns as single units from 
precursor messenger RNA transcripts. However, some long Drosophila melanogaster 
introns contain a cryptic site, known as a recursive splice site (RS-site), that 
enables a multi-step process of intron removal termed recursive splicing. The 
extent to which recursive splicing occurs in other species and its mechanistic 
basis have not been examined. Here we identify highly conserved RS-sites in 
genes expressed in the mammalian brain that encode proteins functioning in 
neuronal development. Moreover, the RS-sites are found in some of the longest 
introns across vertebrates. We find that vertebrate recursive splicing requires 
initial definition of an 'RS-exon' that follows the RS-site. The RS-exon is then 
excluded from the dominant mRNA isoform owing to competition with a 
reconstituted 5' splice site formed at the RS-site after the first splicing 
step. Conversely, the RS-exon is included when preceded by cryptic promoters or 
exons that fail to reconstitute an efficient 5' splice site. Most RS-exons 
contain a premature stop codon such that their inclusion can decrease mRNA 
stability. Thus, by establishing a binary splicing switch, RS-sites demarcate 
different mRNA isoforms emerging from long genes by coupling cryptic elements 
with inclusion of RS-exons.