Enhancement of foreign gene expression by a dicot intron in rice but not in tobacco is correlated with an increased level of mRNA and an efficient splicing of the intron

Abstract

The first intron of castor bean catalase gene, cat-1 was placed in the N-terminal region of the coding sequence of the beta-glucuronidase gene (gusA) and the intron-containing gusA was used with the cauliflower mosaic virus (CaMV) 35S promoter. Using this plasmid, pIG221, the effect of the intron on expression of beta-glucuronidase (GUS) activity was examined in transgenic rice calli and plants (a monocotyledon), and transgenic tobacco plants (a dicotyledon). The intron-containing plasmid increased the level of GUS enzyme activity 10 to 40-fold and 80 to 90-fold compared with the intronless plasmid, pBI221, in transgenic rice protoplasts and transgenic rice tissues, respectively. In contrast, the presence of the intron hardly influenced the expression of the GUS activity in transgenic tobacco plants. Northern blot analysis showed that the catalase intron was efficiently spliced in rice cells while transgenic tobacco plants contained both spliced and unspliced gusA transcripts in equal amounts. Furthermore, the level of the mature gusA transcript in transformed rice calli was greatly increased in the presence of the intron. The catalase intron was removed at the same splice junctions in transgenic rice and tobacco plants. These findings indicate that the stimulating effect of the intron on GUS expression is correlated with an efficient splicing of pre-mRNA and an increased level of mature mRNA.

hpRNA-Mediated Targeting of the Arabidopsis FAD2 Gene Gives Highly Efficient and Stable Silencing

“…Second, the process of intron splicing of the ihpRNA could be responsible for more efficient duplex formation through alignment of the complimentary arms in the spliceosome complex, whereas the complimentary arms of hpRNA would have to hybridize by the less-efficient random but tethered collisions. Third, the mere presence of the intron in the iHP construct may result in increased or morestabilized transcript levels than in the nonintroncontaining hpRNA (Callis et al, 1987;Tanaka et al, 1990). At this stage, we would favor the third explanation for the greater efficiency of the iHP construct to elicit PTGS because Wesley et al (2001) have presented evidence that may discount the first two explanations.…”

Splicing of the Maize Sh1 First Intron Is Essential for Enhancement of Gene Expression, and a T-Rich Motif Increases Expression without Affecting Splicing

“…Plant introns that stimulate expression have been documented in petunia (Petunia hybrida; Dean et al, 1989;Vain et al, 1996), oat (Avena sativa; Bruce and Quail, 1990), rice (Oryza sativa; McElroy et al, 1990;Snowden et al, 1996;Rethmeier et al, 1997), castor bean (Ricinus communis; Tanaka et al, 1990), potato (Solanum tuberosum; Leon et al, 1991;Fu et al, 1995aFu et al, , 1995b, Arabidopsis (Rose and Last, 1997;ChaubetGigot et al, 2001), soybean (Glycine max; Kato et al, 1998), and tobacco (Nicotiana tabacum; Plesse et al, 2001). The approximately 2-to 10-fold range of intron-mediated enhancement usually seen in dicots is much less than increases observed in monocots, which can be more than 100-fold (for review, see Simpson and Filipowicz, 1996).…”

“…The degree of enhancement is usually greater for relatively weak promoters (Callis et al, 1987;Vasil et el., 1987;Mascarenhas et al, 1990;. The magnitude of stimulation also depends on the coding sequences (Sinibaldi and Mettler, 1992;Rethmeier et al, 1997Rethmeier et al, , 1998, the tissue of expression, and physiological conditions (Tanaka et al, 1990;Sinibaldi and Mettler, 1992;Gallie and Young, 1994;Park, 1995a, 1995b;ChaubetGigot et al, 2001;Plesse et al, 2001).…”

“…However, mRNA amounts are not always correlated with increased enzyme activity (Mascarenhas et al, 1990;Tanaka et al, 1990;Bourdon et al, 2001). Tanaka et al(1990) found that decreased splicing efficiency was associated with reduced enzyme activity.…”

“…However, mRNA amounts are not always correlated with increased enzyme activity (Mascarenhas et al, 1990;Tanaka et al, 1990;Bourdon et al, 2001). Tanaka et al(1990) found that decreased splicing efficiency was associated with reduced enzyme activity. There is evidence from animal studies that transcript polyadenylation and transport may be linked to intron splicing (Collis et al, 1990;Huang and Gorman, 1990;Damert et al, 1996).…”