# Gene transcriptions/Boxes/Is

"As in the case of the G box sequence, previous [ribulose-1,5-bisphosphate carboxylase small subunit] RBCS gene expression studies with transgenic plants have failed to demonstrate a requirement for these I box sequences, although mutation of what are likely to be functionally related sequences in CAB genes does affect expression (Gidoni et al., 1989)."
"This [I box] sequence, which is found in most but not all RBCS genes, is also commonly found 5' proximal to the 'TATA' box of most chlorophyll a/b binding protein (CAB) genes (Castresana etal., 1987; Gidoni etal., 1989)."
Experiments "show that site-specific mutations in either G or I, but not GT boxes, in the context of the full rbcS-IA promoter, drastically affect the rbcS-IApromoter-dependent expression of Adh and [β-glucuronidas] GUS reporter genes."
Although "the P3, P6 substitutions alter the conserved 'GATAAG' I box motif, a 'GATA' motif is present in the introduced EcoRV site. This introduced GATA sequence clearly does not serve as a functional I box [...]."
"The sequence targeted by substitution P4 overlaps a sequence showing homology to the 'L' box (consensus 5'-AAATTAACCAA), which is conserved in RBCS upstream sequences of both tomato and tobacco (Giuliano et al., 1988)."
The "I boxes are bound by factor GA-1 (U. Schindler and A. Cashmore, submitted)".

## Consensus sequences

"A third class of conserved DNA sequence, previously referred to as the I box (Giuliano etal., 1988) or sequence 2 (Manzara and Gruissem, 1988) has the consensus 5'-GATAAG."

## Solanum lycopersicum

"The RBCS3A gene of tomato belongs to a small gene family consisting of five members. Although the RBCS1, RBCS2 and RBCS3A promoters contain closely related cis regulatory sequences, the expression patterns of the genes are different. Whereas the RBCS1 and RBCS2 genes are expressed in both leaves and young fruit, the RBCS3A promoter is highly active in leaves, but not in young fruit. This lack of transcription could be due to a mutation in the RBCS3A promoter creating the so-called F-box, a protein binding site located between the activating cis elements, the I-box and G-box."
"In tomato, the RBCS gene family consists of five members at three chromosomal loci (RBCS1, RBCS2, RBCS3A, RBCS3B and RBCS3C). Based on their promoter sequences and expression patterns, the five genes can be divided into two groups (Manzara et al., 1991). The first group consists of the RBCS1, RBCS2 and RBCS3A genes, which contain a conserved pair of I-box (5'-GGATGA-GATAAGA-3') and G-box (5'-CACGTG-3') elements in an identical spatial arrangement (Manzara et al., 1991; Meier et al., 1995)."
It "appears that the RBCS2 promoter and, by extension, the RBCS1 and RBCS3A promoters, are activated by the co-ordinate binding of an I-box binding factor (IBF) and a G-box binding factor (GBF) in leaves, while the promoters are activated by a GBF alone in fruit."
"The I-box motif, 5'-GGATGAGATAAGA-3', or its shorter version 5'-GATAAG-3', has been found in the promoters of a large number of RBCS genes (Giuliano et al., 1988; Manzara and Gruissem, 1988). A related motif (the GATA box) is present in the promoters of the light-regulated chlorophyll a/b binding protein (CAB) genes of different species (Gidoni et al., 1989), and has been shown to be involved in the activation of an Arabidopsis CAB gene by light and by the circadian clock (Anderson and Kay, 1995). I-box and GATA binding factors have been identified in nuclear extracts from tobacco and tomato leaves and cotyledons (Borello et al., 1993; Giuliano et al., 1988; Manzara et al., 1991; Schindler and Cashmore, 1990). The I-box has therefore been suggested to be involved in light-regulated and/or leaf-specific gene expression of photosynthetic genes (Manzara et al., 1991), but to date no I-box binding protein has been cloned from plants."
"The DNA-binding site of LeMYBI was mapped to the motif 5'-GGATGAGATTAGA-3'. This motif, called the I-box, has been found in a large number of RBCS promoters, and has been shown to be necessary for the high-level expression in leaves of the Arabidopsis RBCS-1 A promoter as well as the tomato RBCS2 promoter (Baum et al., 1997; Donald and Cashmore, 1990). The I-box motif is well conserved between the tomato RBCS1, RBCS2 and RBCS3A promoters, where it is paired with a G-box motif in an otherwise unconserved sequence context (Manzara et al., 1993; Meier et al., 1995)."

## Yeast

A "novel protein (LeMYBI) [...] binds specifically to the I-box and activates transcription in yeast and plants. LeMYBI is the first isolated I-box binding protein and is a member of a novel class of myb-like proteins which to date is found exclusively in plants."

## F boxes

"Sequence comparison and protein-DNA interaction studies have shown that the RBCS3A promoter contains a third protein binding site, which overlaps the I-box and the spacer between the I-box and the G-box (Meier et al., 1995). This element, termed F-box, is not present in the RBCS1 and RBCS2 promoters (Meier et al., 1995). It is bound by a fruit-specific DNA-binding activity (FBF), which occupies its site immediately upstream of the G-box and can bind simultaneously with GBF (Meier et al., 1995). Because the binding of FBF correlates with the reduced activity of the RBCS3A promoter in young tomato fruit, FBF has been postulated to be a fruit-specific negative regulator of gene expression (Meier et al., 1995)."
F-box motif is TGATAAG.