Biology subjects

Brieba, L. G.

Publications and source records attributed to Brieba, L. G..

4 recordsLinked to original sources

bioRxiv2026

Plant MutS Homolog 1 is a mismatch-directed nuclease required for organelle genome maintenance

The exceptionally low mutation rates of plant organellar genomes imply the existence of DNA surveillance mechanisms that counteract replication errors and DNA damage. Genetic evidence implicates MutS HOMOLOG 1 (MSH1) as a central component of this pathway, as loss of MSH1 results in the accumulation of point mutations. MSH1 is a unique protein that combines an N-terminal MutS-like mismatch-recognition module with a Cterminal GIYYIG nuclease domain. Here, we show that Arabidopsis thaliana MSH1 (AtMsh1) recognizes mismatches, insertion/deletion loops, and damaged bases within double-stranded DNA and introduces staggered DNA breaks at positions flanking the mismatch or lesion. Given the presence of an active homologous recombination machinery in plant organelles, we hypothesize that these DNA ends may be processed by exonucleases to remove the mismatched or damaged DNA while generating 3' single-stranded DNA substrates suitable for homologous recombination-mediated repair and gene conversion. Together, our findings support a model in which AtMsh1 functions as a minimal mismatch repair system that couples mismatch recognition to DNA incision, providing a potential mechanism for suppressing mutation accumulation and maintaining the remarkable stability of plant organellar genomes

biochemistry↗

bioRxiv2026

Translesion DNA synthesis on pyrimidine dimers by Plant organellar DNA polymerases is metal-dependent

Ultraviolet (UV) radiation generates crosslinked DNA lesions--primarily cyclobutane pyrimidine dimers (CPDs) and [6-4] photoproducts ([6-4] PPs)--that block the progression of replicative DNA polymerases. In plants, these lesions are efficiently removed from nuclear DNA by dedicated repair pathways; however, comparable repair mechanisms are absent in plastids and mitochondria. Consequently, how plant organellar DNA polymerases (POPs) tolerate or bypass UV-induced damage has remained unclear. Here, we show that the two Arabidopsis thaliana organellar polymerases, AtPolIs, possess robust translesion synthesis (TLS) activity across CPDs. Although wild-type enzymes display only limited extension across [6-4] PPs, removal of their exonuclease function dramatically enhances bypass, yielding an efficiency of replication across the [6-4] PP that closely resembles that observed on an undamaged template. This establishes AtPolI as the first known replicative DNA polymerase capable of efficiently bypassing a [6-4] PP. We further demonstrate that TLS across UV photoproducts relies on three unique amino acid insertions within the AtPolI polymerase domain, as deletion of any single insertion abolishes TLS. Notably, Mn{superscript 2} can restore TLS activity in these variants, but only for CPD lesions. Together, these findings identify AtPolIs as the first plant organellar replicases with intrinsic [6-4] PP bypass capability and define the structural features that enable this function. GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=97 SRC="FIGDIR/small/701875v1_ufig1.gif" ALT="Figure 1"> View larger version (15K): org.highwire.dtl.DTLVardef@f497bcorg.highwire.dtl.DTLVardef@15ba2eorg.highwire.dtl.DTLVardef@724e29org.highwire.dtl.DTLVardef@697c8b_HPS_FORMAT_FIGEXP M_FIG C_FIG

biochemistry↗

bioRxiv2026

Pol γ possesses separate metal binding sites for polymerase and strand displacement functions

Accurate replication of mitochondrial genome (mtDNA) integrity, which is essential for cellular metabolism and energy supply, relies primarily on DNA polymerase gamma (Pol {gamma}), Twinkle helicase, and mitochondrial single-stranded DNA binding protein (mtSSB). Twinkle alone exhibits little helicase activity while reports indicate that Pol {gamma} displays from modest to limited unwinding activity. This led us to dissect Pol {gamma} strand displacement activity using structural, biochemical and in silico approaches. Here, we show that human Pol {gamma} carries out robust strand displacement synthesis at physiological concentrations of divalent metal ions which reveals that distinct metal-binding sites can independently regulate DNA synthesis and unwinding activities. We further showed that Pol {gamma} can displace RNA/DNA hybrid with comparable efficiency as DNA/DNA duplex, representing a key implication on RNA primer removal to preserve mtDNA integrity. Our cryo-electron microscopy structures of Pol {gamma} complexed with a template containing downstream dsDNA and an incoming nucleotide revealed the structural mechanism for the strand displacement activity. We identified four conformational states that represent successive stages of DNA unwinding, accompanied by coordinated rearrangement of the downstream DNA and Pol {gamma} elements that mediate strand displacement. This work establishes biochemical and structural mechanisms of Pol {gamma} strand displacement activity, providing fundamental insight into human mitochondrial DNA replication and integrity. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=185 SRC="FIGDIR/small/701366v1_ufig1.gif" ALT="Figure 1"> View larger version (39K): org.highwire.dtl.DTLVardef@1be1dborg.highwire.dtl.DTLVardef@8912c5org.highwire.dtl.DTLVardef@12f5e75org.highwire.dtl.DTLVardef@e25d2a_HPS_FORMAT_FIGEXP M_FIG C_FIG

biochemistry↗

bioRxiv2024

Expansion of the MutS gene family in plants

The MutS gene family is distributed across the tree of life and is involved in recombination, DNA repair, and protein translation. Multiple evolutionary processes have expanded the set of MutS genes in plants relative to other eukaryotes. Here, we investigate the origins and functions of these plant-specific genes. Land plants, green algae, red algae, and glaucophytes share cyanobacterial-like MutS1 and MutS2 genes that presumably were gained via plastid endosymbiotic gene transfer. MutS1 was subsequently lost in some taxa, including seed plants, whereas MutS2 was duplicated in Viridiplantae (i.e., land plants and green algae) with widespread retention of both resulting paralogs. Viridiplantae also have two anciently duplicated copies of the eukaryotic MSH6 gene (i.e., MSH6 and MSH7) and acquired MSH1 via horizontal gene transfer - potentially from a nucleocytovirus. Despite sharing the same name, "plant MSH1" is not directly related to the gene known as MSH1 in some fungi and animals, which may be an ancestral eukaryotic gene acquired via mitochondrial endosymbiosis and subsequently lost in most eukaryotic lineages. There has been substantial progress in understanding the functions of MSH1 and MSH6/MSH7 in plants, but the roles of the cyanobacterial-like MutS1 and MutS2 genes remain uncharacterized. Known functions of bacterial homologs and predicted protein structures, including fusions to diverse nuclease domains, provide hypotheses about potential molecular mechanisms. Because most plant-specific MutS proteins are targeted to the mitochondria and/or plastids, the expansion of this family appears to have played a large role in shaping plant organelle genetics. One-Sentence SummaryPlants are distinguished from other eukaryotes by a functionally diverse complement of MutS proteins gained via a combination of gene duplication, endosymbiotic gene transfer, and horizontal gene transfer.

evolutionary biology↗