Transient expression from cab-m1 and rbcS-m3 promoter sequences is different in mesophyll and bundle sheath cells in maize leaves.

Abstract

Cell-specific and light-regulated expression of the .8-glucuronidase (GUS) reporter gene from maize cab-ml and rbcS-m3 promoter sequences was studied in maize leaf segments by using an in situ transient expression microprojectile bombardment assay. The cab-ml gene is known to be strongly photoregulated and to be expressed almost exclusively in mesophyll cells (MC) but not in bundle sheath cells (BSC).Expression of GUS from a 1026-base-pair 5' promoter fragment of cab-ml is very low in dark-grown leaves; GUS expression is increased about 10-fold upon illumination ofdark-grown leaves. In illuminated leaves, the ratio of GUS expression in MC vs. BSC is about 10:1. The cab-ml region between 868 and 1026 base pairs 5' to the translation start confers strong MC-preferred expression on the remainder of the chimeric gene in illuminated leaves, but a region between -39 and -359 from the translation start is required for photoregulated expression. Transcripts of rbcS-m3 are found in BSC but not in MC and are about double in BSC of greening dark-grown seedlings. In contrast to the behavior of the cab-ml-GUS construct, GUS expression driven by 2.1 kilobase pairs of the rbcS-m3 5' region was about twice as high in MC as in BSC of unilluminated dark-grown maize leaves. The number of BSC, but not MC, expressing GUS nearly doubled upon greening of bombarded etiolated leaves. These data suggest that the 5' region ofrbcS-m3 used here could be responsible for most ofthe light-dependent increase in rbcS-m3 transcripts observed in BSC of greening leaves and that transcriptional or posttranscriptional mechanisms are responsible for the lack of rbcS-m3 transcripts in MC. Although the recent generation of transgenic maize (11) carrying 8-glucuronidase (GUS) reporter gene constructs makes this possible in principle, maize transformation and regeneration procedures are lengthy and thus are not convenient for the analysis of numerous modified forms of genes.We have found that DNA precipitated on tungsten microprojectiles can be delivered into MC and BSC in situ in maize leaf segments and that MC-specific and light-induced GUS expression from the cab-ml promoter1 requires two widely separated sequences 5' to the translation start site. In contrast, GUS expression from the rbcS-m3 promoter is about the same in MC and BSC of 24-hr greened leaves, but expression of the rbcS chimeric gene is promoted by light in BSC. MATERIALS AND METHODSPlant Material. Seeds of maize (Zea mays; FR9cms x FR37; Illinois Foundation Seeds, Champaign, IL) were sown in damp Vermiculite and grown at 300C in darkness. Ten-dayold seedlings were harvested under a dim-green safelight, and their second leaves were used in the transient in situ expression assay described below.Transient in Situ Expression Assay. Four 3.5-cm-long segments of the upper halves of the second leaves of 10-day-old dark-grown maize seedlings, a total area of about 12 cm2, were flattened on 0.8% agar Murashige and Skoog medium (GIBCO) in a 5-cm Petri plate with the lower epidermis facing ...

Tissue-Specific and Developmental Pattern of Expression of the Rice sps1 Gene

“…Computer and visual analysis of the sps1 promoter sequence allowed us to identify a number of DNA motifs potentially involved in the transcriptional regulation of this gene. Putative regulatory elements of two major classes were found to be present in the sps1 promoter: (a) DNA motifs similar to lightresponsive elements (LREs), which are involved in light-and tissue-specific regulation of photosynthesis related genes, such as the I-box (TGGTGNNYAAY-GATAAGG; Argü ello-Astorga and Herrera-Estrella, 1996), G-box (CACGTG; Giuliano et al, 1988), as well as AT-1 like binding sites (AATATTTTTATT; Bansal et al, 1992) and (b) DNA motifs closely related to gibberellin-response elements (GARE) present in ␣-amylase genes such as the Amybox 1 (TAACARA), the O2S box (TATCCAY), and the pyrimidine box (YCTTTTY) (Huang et al, 1990). In addition, a 16-bp sequence (RTGCAATRYWWRAT) present in the pro- Figure 1.…”

“…Promoter region is indicated by an white box, leader region by a hatched box, and exons as black boxes. References for the described sequences are: G-box (Giuliano et al, 1988); AT-1 box (Bansal et al, 1992); I-box (Giuliano et al, 1988); I box core (Gidoni et al, 1989); gibberellin response elements (Huang et al, 1990); and low temperature response elements (Baker et al, 1994). SSCM stands for SPS-SS conserved motif.…”

TRM1, a YY1-like suppressor of rbcS-m3 expression in maize mesophyll cells

“…The expression of GUS reporter genes in MC and BSC was determined by an in situ transient expression assay (7). Four 3.5-cm-long segments from within the upper halves of the second leaves of 10-day-old dark-grown seedlings that had been illuminated for 24 h were flattened side by side on 1.2% agar Murashige and Skoog medium (GIBCO͞ BRL) in 50-mm Petri dishes with the lower epidermis facing upward (9). A suspension of tungsten microprojectiles (1.1 m) on which 5.0 g of reporter gene DNA had been precipitated was shot into maize leaf segments with a PDS-1000 Biolistic apparatus (Bio-Rad).…”

“…We have now identified, within the 3Ј ϩ720 to ϩ957 and the 5Ј Ϫ907 to Ϫ445 bp segments of rbcS-m3 (7,9), three regions that are required for repression. The 3Ј region includes a sequence resembling the repressor form of the binding site for the vertebrate transcription activator-repressor YY1 (Yin Yang 1) (10).…”

Unique and Overlapping Expression Patterns among Members of Photosynthesis-Associated Nuclear Gene Families in Arabidopsis

“…However, highresolution expression profiles in the homologous system are available only for selected members of the RbcS and Lhc families, specific organs, and few species (e.g. Jefferson et al, 1987;Langdale et al, 1988;Silverthorne and Tobin, 1990;Bansal et al, 1992;Meier et al, 1995;Fleming et al, 1996), and are lacking for any of the RbcS and Lhc genes in Arabidopsis.…”