Screening of different embryonic cDNA libraries with the Zmhox1a homeobox (Bellmann and Werr, EMBO J. 11:3367-3374, 1992) led to the isolation of two highly related genes, each containing two homeoboxes. Zmhox2a and Zmhox2b (Zea mays homeobox) encode mRNAs of 6 kb length, and the deduced amino acid sequences exihibit modular proteins of 89% identity. The products of both genes are composed of eight basic NLS-like repeats at the N-terminus, followed by a cysteine rich domain conserved in the Arabidopsis HAT3.1 gene (denoted plant homeodomain finger, Schindler et al., Plant J. 4:137-150, 1993) and two central 159 aa repeats each containing one functional homeodomain. Both homeodomains function independently as DNA-binding motifs. Furthermore eleven proline/glycine rich repeats are found at the C-terminus of the Zmhox2a gene product whereas there are ten in the Zmhox2b protein. Interestingly the central part of the Zmhox2a/2b pair exhibits significant similarity to the Zmhox1a/1b proteins which comprises two N-terminal repeats, the cysteine rich domain and one homeodomain (Fig. 1). Therefore both gene pairs have a common ancestor in evolution.
Northern analysis shows identical transcription patterns for both genes in different maize tissues. High mRNA levels are found in all embryonic stages (Abbe and Stein, Amer. J. Bot. 41, 1954), the plumule and the developing male or female flower, reduced amounts in roots and shoots while only a low transcript level can be detected in dormant embryos and mature leaves.
In situ hybridization experiments (Fig. 2) confirm that the Zmhox2a/b transcripts accumulate in the meristematic regions of all organs analysed. These include the embryonic shoot and root apical meristem, the youngest leaf primordia, the plumule and root tip of the maize seedling and the spikelets of the developing female flower. At the moment transgenic tobacco plants ectopically expressing the Zmhox2a/2b proteins are being regenerated. By this type of gain of function experiments we hope to get hints on the biological function of this double-homeobox gene pair.
Figure 2. In situ hybridization.
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