Mitochondrial RNA Helicases: Key Players in the Regulation of Plant Organellar RNA Splicing and Gene Expression.

Mitochondrial genomes of land plants are large and exhibit a complex mode of gene organization and expression, particularly at the post-transcriptional level. The primary organellar transcripts in plants undergo extensive maturation steps, including endo- and/or exo-nucleolytic cleavage, RNA-base modifications (mostly C-to-U deaminations) and both 'cis'- and 'trans'-splicing events. These essential processing steps rely on the activities of a large set of nuclear-encoded factors. RNA helicases serve as key players in RNA metabolism, participating in the regulation of transcription, mRNA processing and translation. They unwind RNA secondary structures and facilitate the formation of ribonucleoprotein complexes crucial for various stages of gene expression. Furthermore, RNA helicases are involved in RNA metabolism by modulating pre-mRNA maturation, transport and degradation processes. These enzymes are, therefore, pivotal in RNA quality-control mechanisms, ensuring the fidelity and efficiency of RNA processing and turnover in plant mitochondria. This review summarizes the significant roles played by helicases in regulating the highly dynamic processes of mitochondrial transcription, RNA processing and translation in plants. We further discuss recent advancements in understanding how dysregulation of mitochondrial RNA helicases affects the splicing of organellar genes, leading to respiratory dysfunctions, and consequently, altered growth, development and physiology of land plants.

Root Primordium Defective 1 Encodes an Essential PORR Protein Required for the Splicing of Mitochondria Encoded Group II Introns and for Respiratory Complex I Biogenesis.

Cellular respiration involves complex organellar metabolic activities that are pivotal for plant growth and development. Mitochondria contain their own genetic system (mitogenome, mtDNA), which encodes key elements of the respiratory machinery. Plant mtDNAs are notably larger than their counterparts in Animalia, with complex genome organization and gene-expression characteristics. The maturation of the plant mitochondrial transcripts involves extensive RNA editing, trimming and splicing events. These essential processing steps rely on the activities of numerous nuclear-encoded cofactors, which may also play key regulatory roles in mitochondrial biogenesis and function, and hence in plant physiology. Proteins that harbor the Plant Organelle RNA Recognition (PORR) domain are represented in a small gene family in plants. Several PORR members, including WTF1, WTF9 and LEFKOTHEA, are known to act in the splicing of organellar group II introns in angiosperms. The AT4G33495 gene-locus encodes an essential PORR-protein in Arabidopsis, termed as ROOT PRIMORDIUM DEFECTIVE 1 (RPD1). A null mutation of At.RPD1 causes arrest in early embryogenesis, while the missense mutant lines, rpd1.1 and rpd1.2, exhibit a strong impairment in root development and retarded growth phenotypes, especially under high-temperature conditions. Here, we further show that RPD1 functions in the splicing of introns that reside in the coding regions of various complex I (CI) subunits (i.e., nad2, nad4, nad5 and nad7), as well as in the maturation of the ribosomal rps3 pre-RNA in Arabidopsis mitochondria. The altered growth and developmental phenotypes and modified respiration activities are tightly correlated with respiratory chain CI defects in rpd1 mutants.

MSP1 encodes an essential RNA-binding pentatricopeptide repeat factor required for nad1 maturation and complex I biogenesis in Arabidopsis mitochondria.

Mitochondrial biogenesis relies on nuclearly encoded factors, which regulate the expression of the organellar-encoded genes. Pentatricopeptide repeat (PPR) proteins constitute a major gene family in angiosperms that are pivotal in many aspects of mitochondrial (mt)RNA metabolism (e.g. trimming, splicing, or stability). Here, we report the analysis of MITOCHONDRIA STABILITY/PROCESSING PPR FACTOR1 (MSP1, At4g20090), a canonical PPR protein that is necessary for mitochondrial functions and embryo development. Loss-of-function allele of MSP1 leads to seed abortion. Here, we employed an embryo-rescue method for the molecular characterization of msp1 mutants. Our analyses reveal that msp1 embryogenesis fails to proceed beyond the heart/torpedo stage as a consequence of a nad1 pre-RNA processing defect, resulting in the loss of respiratory complex I activity. Functional complementation confirmed that msp1 phenotypes result from a disruption of the MSP1 gene. In Arabidopsis, the maturation of nad1 involves the processing of three RNA fragments, nad1.1, nad1.2, and nad1.3. Based on biochemical analyses and mtRNA profiles of wild-type and msp1 plants, we concluded that MSP1 facilitates the generation of the 3' terminus of nad1.1 transcript, a prerequisite for nad1 exons a-b splicing. Our data substantiate the importance of mtRNA metabolism for the biogenesis of the respiratory system during early plant life.

Group II Intron-Encoded Proteins (IEPs/Maturases) as Key Regulators of Nad1 Expression and Complex I Biogenesis in Land Plant Mitochondria.

Mitochondria are semi-autonomous organelles that produce much of the energy required for cellular metabolism. As descendants of a bacterial symbiont, most mitochondria harbor their own genetic system (mtDNA/mitogenome), with intrinsic machineries for transcription and protein translation. A notable feature of plant mitochondria involves the presence of introns (mostly group II-type) that reside in many organellar genes. The splicing of the mtRNAs relies on the activities of various protein cofactors, which may also link organellar functions with cellular or environmental signals. The splicing of canonical group II introns is aided by an ancient class of RT-like enzymes (IEPs/maturases, MATs) that are encoded by the introns themselves and act specifically on their host introns. The plant organellar introns are degenerated in structure and are generally also missing their cognate intron-encoded proteins. The factors required for plant mtRNA processing are mostly nuclearly-encoded, with the exception of a few degenerated MATs. These are in particular pivotal for the maturation of NADH-dehydrogenase transcripts. In the following review we provide an update on the non-canonical MAT factors in angiosperm mitochondria and summarize the current knowledge of their essential roles in regulating Nad1 expression and complex I (CI) biogenesis during embryogenesis and early plant life.

MISF2 Encodes an Essential Mitochondrial Splicing Cofactor Required for nad2 mRNA Processing and Embryo Development in Arabidopsis thaliana.

Mitochondria play key roles in cellular energy metabolism in eukaryotes. Mitochondria of most organisms contain their own genome and specific transcription and translation machineries. The expression of angiosperm mtDNA involves extensive RNA-processing steps, such as RNA trimming, editing, and the splicing of numerous group II-type introns. Pentatricopeptide repeat (PPR) proteins are key players in plant organelle gene expression and RNA metabolism. In the present analysis, we reveal the function of the MITOCHONDRIAL SPLICING FACTOR 2 gene (MISF2, AT3G22670) and show that it encodes a mitochondria-localized PPR protein that is crucial for early embryo development in Arabidopsis. Molecular characterization of embryo-rescued misf2 plantlets indicates that the splicing of nad2 intron 1, and thus respiratory complex I biogenesis, are strongly compromised. Moreover, the molecular function seems conserved between MISF2 protein in Arabidopsis and its orthologous gene (EMP10) in maize, suggesting that the ancestor of MISF2/EMP10 was recruited to function in nad2 processing before the monocot-dicot divergence ~200 million years ago. These data provide new insights into the function of nuclear-encoded factors in mitochondrial gene expression and respiratory chain biogenesis during plant embryo development.

Why so Complex? The Intricacy of Genome Structure and Gene Expression, Associated with Angiosperm Mitochondria, May Relate to the Regulation of Embryo Quiescence or Dormancy-Intrinsic Blocks to Early Plant Life.

Mitochondria play key roles in cellular-energy metabolism and are vital for plant-life, such as for successful germination and early-seedling establishment. Most mitochondria contain their own genetic system (mtDNA, mitogenome), with an intrinsic protein-synthesis machinery. Although the challenges of maintaining prokaryotic-type structures and functions are common to Eukarya, land plants possess some of the most complex organelle composition of all known organisms. Angiosperms mtDNAs are characteristically the largest and least gene-dense among the eukaryotes. They often contain highly-variable intergenic regions of endogenous or foreign origins and undergo frequent recombination events, which result in different mtDNA configurations, even between closely-related species. The expression of the mitogenome in angiosperms involves extensive mtRNA processing steps, including numerous editing and splicing events. Why do land-plant's mitochondria have to be so complex? The answer to this remains a matter of speculation. We propose that this complexity may have arisen throughout the terrestrialization of plants, as a means to control embryonic mitochondrial functions -a critical adaptive trait to optimize seed germination. The unique characteristics of plant mtDNA may play pivotal roles in the nuclear-regulation of organellar biogenesis and metabolism, possibly to control embryos quiescence or dormancy, essential determinants for the establishment of viable plantlets that can survive post-germination.