Aberrant processing of a Maize GALACTINOL SYNTHASE transcript is caused by heat stress

Abstract

Summary: GALACTINOL SYNTHASE constitutes a highly homologous, small gene family in maize. ZmGOLS2 cDNA probe detects full-length ZmGOLS transcript in dehydration stressed-, and a smaller transcript (ST) in heat stressed-germinating seeds and callus cells. The ST can be detected in seeds imbibed at temperatures above 30 °C, attaining greatest abundance at 40 °C. At 45 °C, the ST is no longer detected and the full-length transcript is again prevalent. Northern blot analysis of poly(A) selected mRNA indicates that the ST is polyadenylated. The ST can be detected by antisense but not by sense RNA probes. Only the five-prime-third of the ZmGOLS2 cDNA is homologous to the ST. However, ribonuclease protection assays (RPA) using a probe to the 5′ portion of ZmGOLS2, led to the conclusion that only ZmGOLS3-, not ZmGOLS2-transcript, is present in heat stressed seeds. The small transcript detected by ZmGOLS2 probe is not derived from ZmGOLS2 but from an unknown, highly homologous gene. Using 3′ RACE, a full-length and a short ZmGOLS3 cDNA were cloned. Sequencing revealed that the short ZmGOLS3 transcript is a fusion of the 5′- and 3′-UTR regions of ZmGOLS3. Comparison with the gene sequence revealed that there are no typical intron–exon junction structures around the deleted fragment of ZmGOLS3. Instead, a five base pair, GC-rich sequence delineates the deletion sites used to form the ZmGOLS3 short transcript. Southern blot analysis using maize genomic DNA as a template confirmed that the aberrant ZmGOLS3 transcript was produced due to aberrant RNA processing and is not due to transcription of a ZmGOLS3 pseudogene.