Exon/intron junctions Exons

Figure 10.13 Phosphoryl-transfer reactions. The figure shows (a) nucleotide polymerization, (b) nucleic acid hydrolysis, (c) first cleavage of an exon-intron junction by group I ribozyme (d) and by a group II ribozyme, (e) strand transfer during transposition and (f) exon ligation during RNA splicing. (From Yang et al., 2006. Copyright 2006, with permission from Elsevier.)...

In the late 1970s and early 1980s, the sequences of many eukaryotic genes became available for study, thus making it possible to conduct a comparative analysis to identify important sequence elements in pre-mRNA. Careful inspection of higher eukaryotic genes revealed several consensus sequences in introns at or near the 5 and 3 splice sites (12, 13). The 5 splice site consensus sequence is G/GURAGU (/ represents the 5 exon-intron junction R depicts a purine the underlined dinucleotide GU is invariant). The 3 splice site is YAG/G (here, / represents the 3 intron-exon junction Y is a pyrimidine the... 

Typing methods involve the design of primer pairs that are able to amplify all alleles at the target HLA locus with the polymorphic sequence motifs situated between the primer sites. Laboratories usually amplify at least exons 2 and 3 of class I genes and exon 2 of class II genes. Some prepare larger amplification products that include exon 1 and/or exon 4 of class I genes. Amplification primers may be located within exons or introns. Primers positioned in introns allow complete analysis of exons and inspection of exon/intron junctions for splice site polymorphisms. Primers must be carefully chosen to attain locus-specific ampfification since... 

While isolating human PS-PLAi cDNA we found a similar cDNA, designated PS-PLAiAC, with four extra bases (GTAC) inserted at the boundary of the exon-intron junction (nt 1122) [23]. The insertion of these four bases caused a frame shift, resulting in the loss of about 80 amino acid residues at the C-terminus (Fig. 2.1). The mRNA having this sequence was found in various human tissues and cell lines [23]. Although it is not clear whether the short form of the enzyme is actually expressed at the protein level, experiments using the recombinant proteins prepared with the baculovirus system revealed the functional difference of the two isoforms (see below) [23]. 

Describe how the discovery that Introns are removed during splicing was made. How are the locations of exon-Intron junctions predicted ... 

Itoga S, Harada S, Nomura F and Nakai T (1998) [Genetic polymorphism of human CYP2E1 new allels detected in exons and exon-intron junctions]. [Japanese]. Nihon Arukoru Yakubutsu Igakkai Zasshi 33 56-64. 

Ohno, K., and Suzuki, K., 1988b, A splicing defect due to an exon-intron junctional mutation results in abnormal P-hexosaminidase a chain mRNAs in Ashkenazi Jewish patients with Tay-Sachs disease, Biochem. Biophys. Res. Commun. 153 463-469. 

C. Splicing errors alter the critical sequence around an intron-exon splice junction. 

Jones et al. (1985) and Yoshimura and Oka (1989) have established the complete exon/intron structure of the rat and mouse (3-casein genes respectively, and, as there is evidence of conservation of exon/ intron junctions in the casein gene family, this work is likely to be directly relevant to bovine (3-Cn. 

The snRNPs are small ribonucleoprotein particles that occur in the nucleus. Each is composed of a small RNA molecule and several characteristic proteins, some of which are common to different snRNPs. Distinct snRNPs recognize and bind to splice junctions and the branch site and are involved in assembling the spliceosome in an ATP-depend-ent manner. They are requisite components of the splicing apparatus, and the RNA components of some of them are probably catalytically active. The RNAs of some snRNPs form hydrogen bonds with sequences within introns and exons to help to juxtapose properly the reacting splice junctions. 

Exons are spliced in a two-stage reaction mediated by snRNP The sequences which are necessary and sufficient to define a splice junction (exon-intron junction) are an invariant GU at the 5 boundary and an invariant AG at the 3 boundary of the intron. The consensus sequence (Padgett et al, 1986) at the exon-intron junctions of eukaryotic pre-mRNA is shown (the subscripts indicate the percentage of pre-mRNA in which the specified base occurs) ... 

---