• What was the problem/issue being addressed?
Every organism faces the challenge to respond favorably to changes in their environment. The blueprint for these responses is their DNA. Cutting-edge genomic technologies show that transcriptional activity predominantly acts on sequences that lack functional annotation. Transcription changes in response to the environment largely occur in non-protein coding DNA regions instead. This suggests that non-coding regions may be part of the response architecture to environmental change. Understanding the role of coding and non-coding transcription dynamics represents a key question in modern biology. By studying the act of transcription, my project gave unprecedented resolution of environmental sensing at the molecular level.
• Why is it important for society?
The cultivars produced by plant breeders have a desired trait, for example more suited to changing environmental conditions. However, the molecular mechanisms of how genomic material helps the adaptation are often unclear. The project drew from cutting-edge studies of transcription dynamics to bridge the gap and elucidate these mechanisms. Thus, the project enabled a potentially more targeted approach to deal with changing environments. There is often a gap of understanding between researchers, the public, industry and policy-makers. To bridge this gap and ensure relevant results are shared with relevant audiences, the project pursued an active communication strategy to make the research visible and openly accessible. Interesting research results was re-written as popular science pieces and published.
• What were the overall objectives?
Objective 1: Develop NET-seq to Arabidopsis to map nascent transcription.
The project aimed to adapt Nascent Elongation Transcript sequencing (NET-seq) to plants (plaNET-seq). With plaNET-seq, we could study the act of transcription in the model plant Arabidopsis thaliana.
Objective 2: Identify environmental responding non-coding RNAs in Arabidopsis with plaNET-seq.
The project used plaNET-seq to map the nascent transcriptome during the response to environmental changes, generating temporal transcription maps of compensatory regulation.
Objective 3: Study the dynamics of transcription in key genomic regions in response to low temperature.
The project looked at specific regulons, important for plant cold-acclimation with a focus on long non-coding RNA function in addition to transcription dynamics (e.g. stalling and release) in response to low temperature.
• What are the conclusions?
The project succeeded in adapting plaNET-seq to Arabidopsis. To achieve the objectives of the action, plaNET-seq was implemented in the early stages of cold-acclimation (3 and 12h of 4°C). Our main conclusions are:
• Transcriptional stalling is greatly affected by cold temperatures. All stages of the transcriptional cycle (initiation, elongation and termination) are affected.
• Generally, the changes in stalling are rapidly adapted between 3h and 12h of treatment to 4°C. This indicates that plants quickly respond and change the chromatin environment and associated transcriptional regulation to cold temperatures.
• With plaNET-seq, close to 10000 novel transcripts were found. This was largely due to their rapid degradation which makes it challenging to detect them with steady-state RNA detection techniques (RNA-seq).
• Many novel non-coding transcripts were found in close proximity to coding genes, predominantly on the antisense strand. plaNET-seq identified for the first time in plants divergent and convergent transcripts. These transcripts are common in yeast and human cells but their function is still largely unknown.
• The characterization of a non-coding transcript (SVALKA) revealed a novel repressive mechanism of gene regulation. SVALKA is transcribed on the antisense strand of the CBF1 gene, a gene important for cold-acclimation in Arabidopsis. Transcription of SVALKA results in RNA Polymerase II collision over the CBF1 gene body, followed by premature termination of the CBF1 mRNA.
