Identification of water-deficit resistance genes in wild almond Prunus scoparia using cDNA-AFLP

• cDNA-AFLP analysis has been used to identify genes responsible for drought tolerance in wild almond P. scoparia identifying twenty differentially expressed transcripts.
• Five differentially expressed genes were annotated to known function genes including AFC2 kinase.
• AFC2 kinase interacts with nuclear mRNA splicing proteins as well as epigenetic response proteins during drought stress.
• The AFC2 derived network has a key regulatory role in P. scoparia drought response.
• Promoter analysis showed that differentially expressed genes harbour binding sites of MYB1 and MYB2 transcription factors which are involved in dehydration response through ABA signalling pathway.

Water deficit is one of the most severe stresses limiting plant growth and yield, inducing the expression of various genes involved in water-deficit tolerance. In this study, we used the differential expression analysis technique cDNA-AFLP (Amplified Restriction Fragment Polymorphism-derived technique for RNA fingerprinting) to identify genes responsible for water stress tolerance in the resistant wild almond Prunus scoparia (Spach). Twenty transcript-derived fragments (TDFs) displayed differential expression between control and stress conditions on polyacrylamide gel. These TDFs were isolated, cloned and sequenced. Sequence alignments using BLAST (Basic Local Alignment Search Tool) showed that several candidate TDFs shared high levels of identity with genes from different species. Furthermore, five differentially expressed genes annotated to known function genes were re-confirmed by quantitative real-time PCR (qRT-PCR), including starch synthase VI, leucine-rich repeat protein, zeaxanthin epoxidase, protein kinase MK5 (AFC2) and 2-deoxyglucose-6-phosphate phosphatase. Network analysis unravelled interaction between protein AFC2 kinase and nuclear RNA splicing proteins (including SR45, SR33, SRZ-22, and RSZP21) which are involved in sugar mediated signalling pathway as well as epigenetics response via histone phosphorelation. Interestingly, promoter analysis showed that differentially expressed genes harbour binding sites of MYB1 and MYB2 transcription factors which are involved in dehydration response through ABA signalling pathway. Our results highlight the importance of starch synthesis, sugar and ABA mediated signalling pathways as well as mRNA splicing and epigenetic response of P. scoparia in resistance to water deficit. These results will be useful for exploring the functions of these multiple signal-inducible genes in order to unveil the relationship and crosstalk between different signalling pathways involved in Prunus resistance to water-deficit. The implication of these results for improving cultivated almonds for drought resistance is also discussed.