Proceedings of the XLVI Italian Society of Agricultural Genetics - SIGA Annual Congress

Giardini Naxos, Italy - 18/21 September, 2002

ISBN 88-900622-3-1

Poster Abstract - 5.16

Monitoring the expression pattern of 624 tomato genes under white/blue light in both wild type and mutants deficient in cryptochrome 1

Giorno F.*, Giliberto L.**, Maugliani A.*, Chiesa M.*, Perrotta G.*

*) ENEA Trisaia Research Center, 75026 Rotondella, Matera

**) ENEA Casaccia Research Center, PO Box 2400, 00100AD Roma, Italy

blue light photoreceptors, expression profiling, microarrays

Cryptochromes (cry) are blue light photoreceptors that mediate many photomorphological responses in land plants such as hypocotyl shortening, cotyledon expansion, induction of anthocyanin synthesis and the control of flowering time. Though Arabidopsis codes for two cryptochromes (Cry1 and Cry2), in angiosperms the CRY gene family seems to span from 2 to 3 elements. Several evidences indicate that Cry1 is the major photoreceptor mediating most of the blue light induced responses under high irradiances, while Cry2 enhances the effect of Cry1 at lower irradiances and promotes flowering.

Despite the relevance of the physiological changes induced by blue light perception, the molecular and functional characterization of cryptochromes has been partially elucidated only in Arabidopsis and tomato. Besides, very little is known about the signalling mechanism and the possible molecular targets of the cryptochromes.

To get further insight the functional interactions between cryptochromes and other molecular species possibly involved in the blue-light responses, we have compared the expression profiling of both tomato wild type and a cry1 deficient mutant probing a tomato cDNA microarray.

In order to characterize the tomato coding sequences for the microarray construction, we have sequenced 1904 clones from a fruit ripening cDNA library. Sequences have been blasted against GenBank, assembled and clustered using the software tools available in GCG Wisconsin Package. As a result 561 non redundant clones representing as many tomato expressed genes has been selected, PCR amplified and spotted over the microarray. The information stored over the microarray has been then further implemented by spotting 63 RT-PCR products corresponding to genes known to be involved in light transduction signals in tomato or in other plant species.

For profiling experiments, tomato seedlings from wild type and cry1 lines have been grown under dim white light for 2 weeks and then exposed to intense blue light for 2hr and 6 hr. At each step part of the plant material has been harvested and RNA extracted. Later, RNA from tomato wild type and cry1 , grown under 2hr and 6 hr of blue light, respectively has been used for synthesis of fluorescent cDNA and compared to the corresponding cDNA from tomato grown in dim white light, by microarray hybridisation. In a second set of experiments the labelled cDNA from wild type has been compared to that of cry1 in conditions of dim white light, 2 hr and 6 hr of blue light treatment, respectively.

Data analysis is currently in progress in order to select genes which possibly show appreciable differences in their expression levels and therefore are candidates to play a role in the mechanism of action of tomato cryptochrome 1.