CTTFMM · Characterization of transcription termination factors in mammalian mitochondria
FP6 — Marie Curie Actions (Human Resources and Mobility)
- Duration
- 2005-05-01 → 2007-04-30
- EU contribution
- €156,883
- Participants
- 1
- Scheme
- EIF
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Results in brief
Final Activity Report Summary - CTTFMM (Characterization of transcription termination factors in mammalian mitochondria)
Human mitochondria contain multiple copies of a small double stranded deoxyribonucleic acid (DNA) genome (mtDNA), of approximately 16.6 kb size, which encodes for two ribosomal ribonucleic acids (RNAs), a set of 22 transfer RNAs and 13 proteins involved in respiration. Gene expression in mitochondria is not self-sufficient since nuclear genes encode all protein components that are required for transcription and translation of the mtDNA-encoded genes as well as all proteins necessary for mtDNA replication. In human cells each of the strands in mtDNA contains one major promoter for transcriptional initiation, the light-strand promoter (LSP) and the heavy-strand promoter (HSP). Transcription from LSP and HSP produces polycistronic precursor RNA encompassing all the genetic information encoded in each of the specific strands. In this Marie Curie financed project, we initiated work to investigate how mitochondrial gene expression was regulated in response to pathological processes and metabolic demands. Proper regulation of mtDNA expression was also likely to require factors that could directly regulate mitochondrial transcription initiation in response to decreased needs for oxidative phosphorylation capacity. We identified a novel family of genes that appeared to play this role in mammalian cells. The mitochondrial transcription termination factor (MTERF) proteins, namely MTERF2, MTERF3, and MTERF4 had the capacity to regulate mitochondrial transcription in vivo. We characterised these factors in a series of unpublished investigations and found that they played a key role in the integration of nuclear and mitochondrial gene expression. The first results of this project were published by Park et al. 2007, 130(2):273-85. In this study, we showed that MTERF3 was a negative regulator of mtDNA transcription initiation. The MTERF3 gene was essential, as homozygous knockout mouse embryos died in mid-gestation. Tissue-specific inactivation of MTERF3 in the heart caused aberrant mtDNA transcription and severe respiratory chain deficiency. MTERF3 bound the mtDNA promoter region and depletion of MTERF3 increased transcription initiation on both mtDNA strands. This increased transcription initiation led to decreased expression of critical promoter-distal tRNA genes, possibly explained by transcriptional collision on the circular mtDNA molecule. MTERF3 was the first example of a mitochondrial protein that acted as a specific repressor of mammalian mtDNA transcription initiation in vivo. In future work, we intended to continue to investigate the molecular and physiological role of the MTERF family of proteins. We also hoped to identify specific substances which could regulate MTERF activity and thereby influence the metabolic activity of mammalian cells.
Data: CORDIS, © European Union
Project objective
Transcription from the mitochondrial (mt) DNA H-strand promoter (HSP) produces a polycistronic precursor RNA encompassing all the genetic information encoded in that strand. Both mt rRNA genes and 12 out of 13 mRNA encoding genes are transcribed from HSP. There is a 20-fold higher expression of the rRNA gene region relative to the downstream mRNA encoding genes which involves an attenuation phenomenon just downstream of the 16S rRNA gene. A central role in this attenuation is played by mTERF, a protein that protects a 28 bp region immediately adjacent to the 3' end of the 16S rRNA gene. The mechanism and regulation of mTERF-dependent transcriptional termination is however still largely unknown.Two works have contributed interesting data, one by the applicant (Asin-Cayuela et al, JBC, 2004) and one by the host laboratory (Falkenberg et al, Nat Genet, 2002). The first one identifies an inactive trimeric form of mTERF, and also presents evidence for a role of mTERF on the regulation of transcription initiation. The second identifies two new mt transcription factors, TFB1M and TFB2M, that can support transcription when combined with the mt RNA polymerase and the mitochondrial transcription factor A. This finding will allow us to investigate the molecular function of mTERF in a defined in vitro system.Moreover, the host laboratory has recently identified an mTERF paralogue in the human genome, TERF2. This protein is ubiquitously expressed, but little is known concerning its structure and function.The aims will be to : 1. Characterize the molecular function of mTERF in a pure in vitro system, in terms of termination activity, interaction with the transcription machinery both in transcription initiation and termination events, mechanism of monomer to trimer transition a nd comparison between HeLa and recombinant mTERF. 2. Investigate the dynamic interactions between mTERF and mtDNA,3. Characterize TERF2.
Original text from CORDIS.
Participants
- KAROLINSKA INSTITUTE · STOCKHOLMCoordinatorSweden
Links
Data: CORDIS, © European Union