Enzymes Involved In Dna Replication And Their Functions PdfBy Malisa O. In and pdf 28.01.2021 at 23:23 8 min read
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- Okazaki fragments
- Proteins Involved in DNA Replication
- Herpes Simplex Viruses: Mechanisms of DNA Replication
Plastids and mitochondria possess their own genomes. Although the replication mechanisms of these organellar genomes remain unclear in photosynthetic eukaryotes, several organelle-localized enzymes related to genome replication, including DNA polymerase, DNA primase, DNA helicase, DNA topoisomerase, single-stranded DNA maintenance protein, DNA ligase, primer removal enzyme, and several DNA recombination-related enzymes, have been identified. In the reference Eudicot plant Arabidopsis thaliana , the replication-related enzymes of plastids and mitochondria are similar because many of them are dual targeted to both organelles, whereas in the red alga Cyanidioschyzon merolae , plastids and mitochondria contain different replication machinery components.
DNA replication employs a large number of proteins and enzymes, each of which plays a critical role during the process. One of the key players is the enzyme DNA polymerase, which adds nucleotides one by one to the growing DNA chain that are complementary to the template strand. The addition of nucleotides requires energy; this energy is obtained from the nucleotides that have three phosphates attached to them, similar to ATP which has three phosphate groups attached.
Plastids and mitochondria possess their own genomes. Although the replication mechanisms of these organellar genomes remain unclear in photosynthetic eukaryotes, several organelle-localized enzymes related to genome replication, including DNA polymerase, DNA primase, DNA helicase, DNA topoisomerase, single-stranded DNA maintenance protein, DNA ligase, primer removal enzyme, and several DNA recombination-related enzymes, have been identified.
In the reference Eudicot plant Arabidopsis thaliana , the replication-related enzymes of plastids and mitochondria are similar because many of them are dual targeted to both organelles, whereas in the red alga Cyanidioschyzon merolae , plastids and mitochondria contain different replication machinery components. The enzymes involved in organellar genome replication in green plants and red algae were derived from different origins, including proteobacterial, cyanobacterial, and eukaryotic lineages.
In the present review, we summarize the available data for enzymes related to organellar genome replication in green plants and red algae. In addition, based on the type and distribution of replication enzymes in photosynthetic eukaryotes, we discuss the transitional history of replication enzymes in the organelles of plants. Plastids and mitochondria are semi-autonomous organelles that contain their own genomes, encoding the genes necessary to perform their respective metabolic functions.
In contrast, plant organellar genomes do not encode these replicative proteins, and are instead replicated by nucleus-encoded enzymes that are transported to the organelles. Studies of bacterial replication enzymes Langston et al. All of the enzymes involved in mitochondrial DNA replication in vertebrates have been identified Arnold et al.
A number of other replication-related enzymes, including topoisomerases 1 and 3a, SSB, ligase 3, and RNase H1, have also been identified in human mitochondria. Several of the replicative enzymes found in bacteria and animal mitochondria are also encoded by plant nuclear genomes. In addition to these common enzymes, a number of plant-specific enzymes for DNA replication and recombination have recently been identified, and their subcellular localization has been examined in both plants and algae.
In this article, we summarize the current knowledge on enzymes related to organellar replication in photosynthetic eukaryotes and also discuss the evolution of these replication-related enzymes based on their distribution in photosynthetic eukaryotes. DNA replication activity was first detected in isolated organelles from plants, yeasts, and animals in the late s Wintersberger, ; Parsons and Simpson, ; Spencer and Whitfeld, ; Tewari and Wildman, In the following decade, DNA polymerases were purified from isolated chloroplasts and mitochondria of various photosynthetic eukaryotes summarized in Moriyama and Sato, Sakai et al.
This finding led to the identification of a gene s encoding a DNA polymerase with distant homology to E. The identified DNA polymerase was first isolated from plastids of rice, and its localization was confirmed by immunoblot analysis of isolated plastids Kimura et al.
We also identified this type of DNA polymerase in algae and ciliates Moriyama et al. In addition, red algae were found to encode a DNA polymerase with high homology to E. Therefore, we proposed that this type of organellar DNA polymerase be named POP plant and protist organellar DNA polymerase , because the genes encoding the polymerases are present in both photosynthetic eukaryotes and protists.
Reproduced from Moriyama et al. POPs also show divalent metal ion-dependent activity, and the optimal MgCl 2 concentration for their activity is 2. DNA polymerase enzymes bind to and dissociate from template DNA repeatedly during the replication or repair process. The number of synthesized nucleotides added by the DNA polymerase per one binding event is defined as processivity. In comparison, E. In contrast, POPs show high processivity as a single subunit enzyme, and to our knowledge, no accessory proteins associated with POP have been identified.
PAA was originally identified as an inhibitor of viral DNA polymerases and reverse transcriptases, and functions by interacting with pyrophosphate binding sites, leading to an alternative reaction pathway Leinbach et al. This exonuclease activity was shown in rice Takeuchi et al. The spatial expression patterns of POPs were analyzed in A. The analysis demonstrated that AtPOP1 is mainly expressed in rosette leaves, whereas AtPOP2 is predominantly found in the meristems of roots and shoots.
The unicellular red alga Cyanidioschyzon merolae contains a single plastid and mitochondrion Matsuzaki et al. In synchronous cultures of Cyanidioschyzon merolae established using light—dark cycles Suzuki et al. Replication of the mitochondrial genome appears to be at least partially synchronized with the cell cycle, as mitochondrial DNA begins to replicate from the light phase, and reaches a two-fold increase at or near the M-phase.
In contrast, plastid DNA increases gradually throughout the entire cell cycle, even after cell division is complete Moriyama et al. However, the protein level of POP remains nearly unchanged throughout the cell cycle, with only small increases and decreases occurring during the light and dark phases, respectively. POP mutants of A. The A. Additionally, the Atpop2 mutant displayed high sensitivity to ciprofloxacin, an inducer of DNA double-strand breaks. TWINKLE T7 gp4-like protein with intramitochondrial nucleoid localization , which is a homolog of the T7 phage gp4 protein with primase and helicase activities, was originally reported to function as a hexameric DNA helicase in human mitochondria Spelbrink et al.
Dual-targeted enzymes to the mitochondria and chloroplasts of plants are summarized in the review by Carrie and Small Red algae and diatoms have a plastid-encoded DnaB helicase and a nucleus-encoded DnaG primase. We also confirmed the plastid-localization of DnaG in the red alga Porphyridium purpureum.
In addition to gyrases, plant organelles contain A-type topoisomerase I, which is a homolog of bacterial topoisomerase I TopA. To search for mitochondrial topoisomerases, we examined the subcellular localization of topoisomerases encoded in the Cyanidioschyzon merolae genome, and showed that a homolog of eukaryotic TOP2 is targeted to mitochondria.
To date, organellar localization of eukaryotic TOP2 has not been reported in plants. In Cyanidioschyzon merolae , the gyrase specific inhibitor nalidixic acid arrests not only replication of the plastid genome, but also that of the mitochondrial and nuclear genomes Itoh et al.
The localization results of gyrases in Cyanidioschyzon merolae suggest that defective plastid replication leads to the arrest of mitochondrial and nuclear replication by a yet unknown mechanism. Four DNA ligases have been identified in the A. AtLIG1 is expressed in all tissues of A. However, it has been noted that AtLIG6 has a putative plastid-targeting peptide at the N -terminus and might therefore be targeted to plastids Sunderland et al.
Cyanidioschyzon merolae has a single gene encoding DNA ligase. Cyanidioschyzon merolae DNA ligase 1 CmLIG1 has two methionine residues in its N -terminal region and is targeted to both mitochondria and plastids when the transcript is translated from the first and second initiation codons Moriyama et al. Therefore, CmLIG1 appears to have triple localization in plastids, mitochondria, and the nucleus. AtSSB1 is localized to mitochondria, but was also reported to be localized to chloroplasts in the review by Cupp and Nielsen PDF motifs are conserved only in green plants, including Chlamydomonas reinhardtii.
In our analysis, the SSB of Cyanidioschyzon merolae is localized only in the mitochondrion, unlike that of A. We performed the localization analysis using a construct starting from the second methionine codon or starting from the ATA codon located upstream of the first methionine codon; however, none of the constructs showed plastid localization.
We also examined the organellar localization of RPAs in Cyanidioschyzon merolae , and even though they have no extension sequence at the N -terminus, they were localized to the nucleus. Based on these findings, the plastidial SSB in red algae remains unidentified. In contrast, RNaseH1 performs this role in human mitochondria Kasiviswanathan et al. Cyanidioschyzon merolae has a gene with high sequence homology to bacterial PolI Moriyama et al.
Phylogenetic analyses of bacterial-type replicative enzymes have been performed Moriyama et al. Phylogenetic trees of enzymes related to organellar genome replication. Simplified phylogenetic trees A—D. Modified from Moriyama et al. DnaB and DnaG are conserved only in red algae. The retention of SSBs is highly variable in photosynthetic eukaryotes.
Bacterial-type SSB proteins are conserved in land plants and Cyanidioschyzon merolae , whereas OSB proteins are conserved among land plants, including A. According to this classification, Physcomitrella patens and K. Conservation of origin-binding protein ODB is more limited, as only land plants have this protein.
Therefore, all photosynthetic eukaryotes contain proteobacteria-derived PolI. The observed distribution of enzymes that play key roles in replication indicates that they are essentially conserved in all plants and algae. In contrast, because recombination-related enzymes and SSBs are non-uniformly distributed among plants and algae, these enzymes are considered to exhibit high plasticity during evolution. List of replication-related enzymes possibly localized to plastids or mitochondria in photosynthetic eukaryotes.
In red algae, most replication enzymes in the ancestor of photosynthetic eukaryotes are found in present-day species. Proposed model for the exchange of organellar replication enzymes during the evolution of photosynthetic eukaryotes. In the past decade, most enzymes related to plastid and mitochondrial DNA replication in plants and algae have been identified.
These studies have revealed that the core enzymes and components involved replication are identical in the plastids and mitochondria of land plants. In contrast, SSBs and recombination-related enzymes are not universally conserved in the green lineage, suggesting that these enzymes are possibly susceptible to exchange or loss during evolution, leading to the acquisition or creation of species-specific enzymes.
Unlike the green lineage, red algae contain different replicative protein profiles in plastids and mitochondria.
Red algal plastids contain numerous replication proteins that originated from cyanobacteria Moriyama et al. To date, a number of organelle-localized enzymes have been identified. However, biochemical data are lacking for the majority of organellar replication enzymes in plants.
The role of an enzyme predicted by homology searches against known enzymes might differ from its actual function or properties. The regulatory mechanisms controlling the initiation of plant organellar genome replication and the number of organellar DNA copies remains to be explored. Recently, chloroplast DNA replication was shown to be regulated by the cellular redox state in the green alga Chlamydomonas reinhardtii Kabeya and Miyagishima, Specifically, chloroplast DNA replication was activated and inactivated by the addition of reducing and oxidative agents, respectively, in both in vivo and in vitro assays.
Light-dependent genome replication was also reported in cyanobacteria, in which DCMU [3- 3,4-dichlorophenyl -1,1-dimethylurea], an inhibitor of electron transport between the PSII complex and plastoquinone pool, inhibits DNA replication initiation, and DBMIB 2,5-dibromoisopropylmethyl-p-benzoquinone , an inhibitor of electron transport between plastoquinone and cytochrome b 6 f complex, inhibits the initiation and elongation of replication Watanabe et al.
Thus, the light-mediated replication of plastid DNA in algae may have originated from cyanobacteria. However, organellar replication in land plants and multicellular plants appears to be regulated by other mechanisms. In land plants, the replication of organellar genomes is restricted to meristematic tissues, and is not associated with the cycle or organellar division Hashimoto and Possingham, ; Fujie et al.
These findings suggest that land plants have more complex regulatory mechanisms controlling the replication of organellar genomes than those operating in algae. The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest. National Center for Biotechnology Information , U.
Journal List Front Plant Sci v. Front Plant Sci. Published online Sep
Proteins Involved in DNA Replication
Okazaki fragments are short sequences of DNA nucleotides approximately to base pairs long in eukaryotes which are synthesized discontinuously and later linked together by the enzyme DNA ligase to create the lagging strand during DNA replication. During DNA replication, the double helix is unwound and the complementary strands are separated by the enzyme DNA helicase , creating what is known as the DNA replication fork. This causes periodic breaks in the process of creating the lagging strand. The primase and polymerase move in the opposite direction of the fork, so the enzymes must repeatedly stop and start again while the DNA helicase breaks the strands apart. Once the fragments are made, DNA ligase connects them into a single, continuous strand.
The enzymes involved in organellar genome replication in green plants and red algae were derived from different origins, including.
Herpes Simplex Viruses: Mechanisms of DNA Replication
An illustration to show replication of the leading and lagging strands of DNA. Image credit: Genome Research Limited. Each genome contains all of the information needed to build that organism and allow it to grow and develop.
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DNA polymerase δ replicates the leading strand, while DNA polymerase ϵ synthesizes the lagging strand. DNA polymerase α catalyzes priming of both the strands. DNA polymerases ξ, η, τ, and k are all nuclear DNA repair enzymes. DNA polymerase γ is found in mitochondria and catalyzes replication of mtDNA.