AbstractMajor changes in expression occur in Arabidopsis in response to cold. It is now clear that genome-wide changes in alternative splicing (AS) also occur in Arabidopsis during the cold-response and many of the genes which undergo cold-induced AS have been linked to roles in either the regulation of the cold-response or regulation of AS. Mutations in splicing factors in Arabidopsis, such as STA1 and SRL1 have been shown to lead to both changes in AS and changes in cold-sensitivity/tolerance, suggesting an important the role for AS in regulating the cold-response. Less is known about the effect of cold on AS in barley or how AS impacts the barley cold-response. There are only a few studied examples of cold-induced AS changes in barley genes, although this is rapidly changing due to both the publication of the barley genome and next generation sequencing of the transcriptome. To investigate AS in the barley cold response, 11 barley genes with cold-induced AS changes were identified and the AS change was analyzed in detail. The barley genes FRY2 and SUA change AS after 30 minutes exposure to cold and are both genes have been linked to roles in regulating AS, indicating a complex role for AS in the earliest stages in the cold-response. The Serine Arginine (SR)-Rich protein genes have been shown to change AS and affect AS under stress conditions in Arabidopsis, rice and other plant species but little is known about the SR protein genes in barley or how the genes change splicing/expression in response to cold. The 16 members of the barley SR protein gene family were identified and analyzed for cold-induced expression changes using available microarray and RNA-Seq data. The HvRS41 gene showed a >2 fold increase in expression after 3h exposure to 6°C in a cold-based microarray experiment. A cold-based microarray experiment in Arabidopsis showed a similar cold-induced expression of the AtRS40 gene, a RS-type SR protein gene with high homology to HvRS41. The cold-induced expression of HvRS41 and AtRS40 indicate a role for the RS-type SR protein genes in the cold response. The RS-type SR proteins form a splicing complex with FRY2 which could potentially be regulated through both AS and expression change during the cold-response. The Barley SR Proteins were dived into six sub-groups previously established for plant SR protein genes. Five out of the six sub-groups of the barley SR protein genes contained AS which could be validated through RT-PCR based methods. The SR-type SR protein genes contained was shown to contain three genes (HvSR34, HvSR30a and HvSR34) within barley. All 3 barley SR-type protein genes showed AS change in response to low temperatures, indicating a role for the barley SR-type SR protein genes in regulating AS during the cold-response. The role for SR-type SR protein genes in regulating AS was tested through a creation of a barley transgenic line over-expressing gene the HvSR34. The HvSR34 overexpression lines are in the process of being tested for changes in AS and cold tolerance.
|Date of Award||2016|
|Sponsors||The James Hutton Institute & Biotechnology and Biological Sciences Research Council|
|Supervisor||John Brown (Supervisor) & Craig G. Simpson (Supervisor)|
- Alternative splicing
The Effect of Low Temperature on Alternative Splicing in Barley
Raeside, A. (Author). 2016
Student thesis: Doctoral Thesis › Doctor of Philosophy