We are searching data for your request:
Upon completion, a link will appear to access the found materials.
The nucleus is the largest and most prominent of a cell’s organelles (Figure 3.19). The nucleus is generally considered the control center of the cell because it stores all of the genetic instructions for manufacturing proteins. Interestingly, some cells in the body, such as muscle cells, contain more than one nucleus (Figure 3.20), which is known as multinucleated. Other cells, such as mammalian red blood cells (RBCs), do not contain nuclei at all. RBCs eject their nuclei as they mature, making space for the large numbers of hemoglobin molecules that carry oxygen throughout the body (Figure 3.21). Without nuclei, the life span of RBCs is short, and so the body must produce new ones constantly.
Figure 3.19. The NucleusThe nucleus is the control center of the cell. The nucleus of living cells contains the genetic material that determines the entire structure and function of that cell.
Figure 3.20. Multinucleate Muscle CellFUnlike cardiac muscle cells and smooth muscle cells, which have a single nucleus, a skeletal muscle cell contains many nuclei, and is referred to as “multinucleated.” These muscle cells are long and fibrous (often referred to as muscle fibers). During development, many smaller cells fuse to form a mature muscle fiber. The nuclei of the fused cells are conserved in the mature cell, thus imparting a multinucleate characteristic to mature muscle cells. LM × 104.3. (Micrograph provided by the Regents of University of Michigan Medical School © 2012)
Figure 3.21. Red Blood Cell Extruding Its NucleusMature red blood cells lack a nucleus. As they mature, erythroblasts extrude their nucleus, making room for more hemoglobin. The two panels here show an erythroblast before and after ejecting its nucleus, respectively. (credit: modification of micrograph provided by the Regents of University of Michigan Medical School © 2012)
Inside the nucleus lies the blueprint that dictates everything a cell will do and all of the products it will make. This information is stored within DNA. The nucleus sends “commands” to the cell via molecular messengers that translate the information from DNA. Each cell in your body (with the exception of germ cells) contains the complete set of your DNA. When a cell divides, the DNA must be duplicated so that the each new cell receives a full complement of DNA. The following section will explore the structure of the nucleus and its contents, as well as the process of DNA replication.
Organization of the Nucleus and Its DNA
Like most other cellular organelles, the nucleus is surrounded by a membrane called the nuclear envelope. This membranous covering consists of two adjacent lipid bilayers with a thin fluid space in between them. Spanning these two bilayers are nuclear pores. A nuclear pore is a tiny passageway for the passage of proteins, RNA, and solutes between the nucleus and the cytoplasm. Proteins called pore complexes lining the nuclear pores regulate the passage of materials into and out of the nucleus. Inside the nuclear envelope is a gel-like nucleoplasm with solutes that include the building blocks of nucleic acids. There also can be a dark-staining mass often visible under a simple light microscope, called anucleolus (plural = nucleoli). The nucleolus is a region of the nucleus that is responsible for manufacturing the RNA necessary for construction of ribosomes. Once synthesized, newly made ribosomal subunits exit the cell’s nucleus through the nuclear pores. The genetic instructions that are used to build and maintain an organism are arranged in an orderly manner in strands of DNA. Within the nucleus are threads of chromatin composed of DNA and associated proteins (Figure 3.22). Along the chromatin threads, the DNA is wrapped around a set of histone proteins. Anucleosome is a single, wrapped DNA-histone complex. Multiple nucleosomes along the entire molecule of DNA appear like a beaded necklace, in which the string is the DNA and the beads are the associated histones. When a cell is in the process of division, the chromatin condenses into chromosomes, so that the DNA can be safely transported to the “daughter cells.” The chromosome is composed of DNA and proteins; it is the condensed form of chromatin. It is estimated that humans have almost 22,000 genes distributed on 46 chromosomes.
Figure 3.22. DNA MacrostructureStrands of DNA are wrapped around supporting histones. These proteins are increasingly bundled and condensed into chromatin, which is packed tightly into chromosomes when the cell is ready to divide.
Viral and cellular interactions during adenovirus DNA replication
Adenoviruses represent ubiquitous and clinically significant human pathogens, gene-delivery vectors, and oncolytic agents. The study of adenovirus-infected cells has long been used as an excellent model to investigate fundamental aspects of both DNA virus infection and cellular biology. While many key details supporting a well-established model of adenovirus replication have been elucidated over a period spanning several decades, more recent findings suggest that we have only started to appreciate the complex interplay between viral genome replication and cellular processes. Here, we present a concise overview of adenovirus DNA replication, including the biochemical process of replication, the spatial organization of replication within the host cell nucleus, and insights into the complex plethora of virus-host interactions that influence viral genome replication. Finally, we identify emerging areas of research relating to the replication of adenovirus genomes.
Keywords: DNA adenovirus liquid-liquid phase separation viral replication compartment virus.
© 2019 Federation of European Biochemical Societies.
Figure 1.. Overview of HAdV replication cycle.
Figure 1.. Overview of HAdV replication cycle.
Virus entry and import of viral genomes into…
Figure 2.. Schematic representation of the Ad5…
Figure 2.. Schematic representation of the Ad5 genome.
The organization of genes within the central…
Figure 3.. Replication of the HAdV genome…
Figure 3.. Replication of the HAdV genome by strand displacement.
Following replication initiation, the viral…
Figure 4.. Viral replication compartments are reorganized…
Figure 4.. Viral replication compartments are reorganized during the late stage of infection.
In order for an organism to grow, develop, and maintain its health, cells must replicated themselves by dividing to produce two new daughter cells, each with the full complement of DNA as found in the original cell. Billions of new cells are produced in an adult human every day. There are a few cell types in the body do not divide, including nerve cells, skeletal muscle fibers, and cardiac muscle cells. The division time of different cell types varies. Epithelial cells of the skin and gastrointestinal lining, for instance, divide very frequently to replace those that are constantly being rubbed off of the surface by friction.
A DNA molecule is made of two strands that “complement” each other: the molecules that compose the strands fit together and bind to each other, creating a double-stranded molecule that looks much like a long, twisted ladder. Each side rail of the DNA ladder is composed of alternating sugar and phosphate groups (Figure 4).
Figure 4. Molecular Structure of DNA. The DNA double helix is composed of two complementary strands. The strands are bonded together via their nitrogenous base pairs using hydrogen bonds.
The two sides of the ladder are not identical, but are complementary. These two backbones are bonded to each other across pairs of protruding bases, each bonded pair forming one “rung,” or cross member. The four DNA bases are adenine (A), thymine (T), cytosine (C), and guanine (G). Because of their shape and charge, the two bases that compose a pair always bond together. Adenine always binds with thymine, and cytosine always binds with guanine. The particular sequence of bases along the DNA molecule determines the genetic code. Therefore, if the two complementary strands of DNA were pulled apart, you could infer the order of the bases in one strand from the bases in the other, complementary strand. For example, if one strand has a region with the sequence AGTGCCT, then the sequence of the complementary strand would be TCACGGA.
DNA replication is the copying of DNA that occurs before cell division can take place. After a great deal of debate and experimentation, the general method of DNA replication was deduced in 1958 by two scientists in California, Matthew Meselson and Franklin Stahl. This method is illustrated in Figure 5 and described below.
Figure 5. DNA Replication. DNA replication faithfully duplicates the entire genome of the cell. During DNA replication, a number of different enzymes work together to pull apart the two strands so each strand can be used as a template to synthesize new complementary strands. The two new daughter DNA molecules each contain one pre-existing strand and one newly synthesized strand. Thus, DNA replication is said to be “semiconservative.”
Stage 1: Initiation
The two complementary strands are separated, much like unzipping a zipper. Special enzymes, including helicase, untwist and separate the two strands of DNA.
Stage 2: Elongation
Each strand becomes a template along which a new complementary strand is built. DNA polymerase brings in the correct bases to complement the template strand, synthesizing a new strand base by base. A DNA polymerase is an enzyme that adds free nucleotides to the end of a chain of DNA, making a new double strand. This growing strand continues to be built until it has fully complemented the template strand.
Stage 3: Termination
Once the two original strands are bound to their own, finished, complementary strands, DNA replication is stopped and the two new identical DNA molecules are complete. Each new DNA molecule contains one strand from the original molecule and one newly synthesized strand. The term for this mode of replication is “semiconservative,” because half of the original DNA molecule is conserved in each new DNA molecule. This process continues until the cell’s entire genome, the entire complement of an organism’s DNA, is replicated. As you might imagine, it is very important that DNA replication take place precisely so that new cells in the body contain the exact same genetic material as their parent cells. Mistakes made during DNA replication, such as the accidental addition of an inappropriate nucleotide, have the potential to render a gene dysfunctional or useless. Fortunately, there are mechanisms in place to minimize such mistakes. A DNA proofreading process enlists the help of special enzymes that scan the newly synthesized molecule for mistakes and corrects them. Once the process of DNA replication is complete, the cell is ready to divide. You will explore the process of cell division later in the chapter.
Watch this video to learn about DNA replication. DNA replication proceeds simultaneously at several sites on the same molecule. What separates the base pair at the start of DNA replication?
3.4: The Nucleus and DNA Replication - Biology
An excellent question. Mitochondria divide by simple fission, splitting in two just as bacterial cells do, and although the DNA replication strategies are a little different, forming displacement or D-loop structures, they partition their circular DNA in much the same way as do bacteria. Mitochondrial reproduction is not autonomous (self-governed), however, as is bacterial reproduction. Most of the components required for mitochondrial division are encoded as genes within the eukaryotic (host) nucleus and translated into proteins by the cytoplasmic ribosomes of the host cell. Mitochondrial replication is thus impossible without nuclear participation, and mitochondria cannot be grown in a cell-free culture. A tight control over mitochondrial division is essential to prevent uncontrolled mitochondrial replication, which could easily lead to destruction of the host cell. This provides an elegant illustration of the co-evolution between the mitochondria and their hosts in the evolution of the eukaryota.
Mitochondria and chloroplasts divide by fission, much like bacteria. When the cell divides, the mito and chloro are distributed to the daughter cells.
Most of the proteins in the mito and chloro are encoded by the nuclear genome and they are imported. They are translated on ribosomes in the cytoplasm. The newly formed protein has a sequence of amino acids at the N-terminus that acts as an import signal - it is recognized and bound by the import machinery on the membrane of the mito or chloro and the protein is pulled inside.
Wow, what a good question! I never thought to ask about that when I took my biology classes in college. Doing some research, I found that this is an area scientists don't know much about. I'm not at all knowledgeable on the subject, so I got a friend of mine to help me out (Ed Lowry, a graduate student in the Department of Ecology, Evolution and Marine Biology at UC Santa Barbara). Remember that chloroplasts and mitochondria are known as organelles (another word for membrane-bound bodies within a cell), and the cytosol is the liquid within the cytoplasm, or the interior of the cell. Here's what Ed had to say:
Some old texts of mine bring a few interesting bits to light. 3&4 are the most pertinent ones. All quotes are from Ch. 7 of Molecular Biology of the Cell, by Alberts and his buddies:
1) Mitochondria in the cell are not strictly individuals. They are "remarkably plastic organelles, constantly changing their shape, even fusing with one another and then separating again."
2) Mitochondria and chloroplasts are dependent for the most part on proteins synthesized from nuclear DNA and imported into the organelle. Some proteins are encoded by organelle DNA and synthesized in the organelle. Interestingly, "no protein is known to be exported from mitochondria or chloroplasts to the cytosol."
3) A class of yeast mutants called "cytoplasmic petite mutants" entirely lack DNA in their mitochondria. "Although petite mutants cannot synthesize proteins in their mitochondria, and therefore cannot make mitochondria that produce ATP, they nevertheless contain mitochondria that have a normal outer and an inner membrane with poorly developed cristae [the folds in the membrane]. Such mutants dramatically demonstrate the overwhelming importance of the nucleus in biogenesis. They also show that an organelle that [here's the important part!] divides by fission can replicate indefinitely in the cytoplasm of proliferating eukaryotic cells even in the complete absence of its own genome."
4) "Overall control [of organelle replication] clearly resides in the nucleus. the nucleus must regulate the number of mitochondria and chloroplasts in the cell according to need. Although these regulatory aspects are crucial to our understanding of eukaryotic cells, we know relatively little about them." Well, shoot.
For most cell types there is what is called a "restriction point" in the cell cycle. Prior to this point the cells might maintain a sort of status quo if, for example, the environment is unfavorable for growth. Past this point, an internal change takes place which commits the cell to replicate its DNA and divide.
A signaler called "S-phase activator" (I guess people with lots of imagination become screenwriters or clothing designers) appears in the cytoplasm prior to DNA replication. (the book says it may be a group of molecules and not a single one, but it doesn't specify their identities). The major control molecules have been related to a class of genes termed "cdc" genes, for "cell-division cycle", of which there are more than twenty.
DNA Structure: Gumdrop Modeling
In this 3-part lab, students will get an up-close and personal look at DNA, including its structure, how that structure is important for its replication, and how it is packaged and regulated.
Download the labs!
Student Advanced Version
- Both versions of the lab use a ruler to make basic measurements, which should be understandable for almost any age.
- The advanced lab has a few thought / concept questions which are likely difficult for those below high school science levels.
- DNA is made of strings of nucleotides. A nucleotide is a chemical molecule composed of one phosphate group, one sugar ring, and one nitrogen-containing base.
- DNA has 4 types of bases: Adenine, Thymine, Cytosine, and Guanine (A, T, C, and G). These bases have strict binding rules, as A only bonds with T (and vice versa), and C only bonds with G. This is important for DNA replication to work.
- DNA is carefully packaged in the nucleus to compact it, protect it, and control which parts of the DNA are turned on and off in different cells.
Part 1 & 2 (per group of 2-4 students) – DNA Models
- 1 box DOTS gumdrop candy (5 colors per box) per 3 groups
- 1 box Crows licorice candy per 3-4 groups
- 1 box Twizzlers Bites (or similar*) per 3-4 groups
- 35 Toothpicks (flat, not round, if possible)
Part 3 – Strawberry DNA Extraction (also done in groups)
- Measuring cup
- Measuring spoons
- Rubbing alcohol
- Strawberries (3 per group)
- Dish detergent
- Cheesecloth or coffee filter
- Tall cup / glass
- Sealable sandwich bags
- Test tubes (optional)
Part 4 – DNA Compaction (per group)
- A ruler
- A calculator
- 2 meters of sewing thread
- 2 pieces of white string or yarn, with colored patches (location is important (one towards the end, and one roughly centered), but you don’t need to label them)
- 2 strips of Scotch tape, wrapped into a circle (sticky side out)
*Other pliable, but firm, candies can be substituted, such as “Circus Peanuts” or small caramels
Completion stage of replication: DNA Polymerase I
Once DNA Polymerase III pairs the majority of complementary D-nucleotides on the developing leading and lagging strand, the RNA primers are still present. Another DNA Polymerase enzyme, DNA Polymerase I reads both the leading and lagging strands from 5’ to 3’ of the parental strands replacing the R-nucleotides with D-nucleotides. While the leading strand is synthesized continuously, there is still an RNA primer at the origin of the replication bubble. It is also replaced by DNA Polymerase I.
Completion stage of replication: Ligase
After DNA Polymerase I replaces the RNA primers, there is no phosphodiester linkage between Okazaki fragments. Another enzyme, ligase, travels down the lagging strand creating a phosphodiester linkage connecting the Okazaki fragments.
Ligase binds Okazaki fragments together.
Completion stage of replication: Telomerase
In prokaryotes, once ligase is finished connecting the Okazaki fragments, the DNA has been successfully replicated. This is due to the circular shape of their DNA. However, eukaryotic DNA is linear. In the lagging strand this poses a problem. At the end of the chromosomes, known as the telomeres, there is nowhere for the DNA Polymerase III to connect due to the absence of an RNA primer. If these D-nucleotides at the telomeres were left unsynthesized , the DNA strands would eventually become shorter with every replication. It is thought that aging is a product of malfunctioning telomerase.
Telomerase extends the ends (telomeres) of the lagging strand.
Telomerase is an enzyme that attaches to the unsynthesized end of the lagging strand and catalyzes the synthesis of DNA from its own RNA template, akin to reverse engineering. At one end of the telomerase a few R-nucleotides combine with the end of the unreplicated strand. Telomerase then adds D-nucleotides to the end of the lagging strand.
The opposite ends of telomerase are identical. The opposite side of the telomerase (if you read left to right) ends with the same R-nucleotides (AUU). Telomerase uses this phenomenon to detach once the lagging strand has been extended and then reattach back to the end of the extended lagging strand. The reattached telomerase replicates another exact copy to the lagging strand. This process happens several times and the telomerase eventually is removed.
Once telomerase lengthens the parental lagging strand, primase attaches synthesizing an RNA primer, which allows DNA Polymerase III to replicate the remaining D-nucleotides on the unreplicated strand. Ligase fuses the last remaining segment by creating the final phosphodiester linkage. This results in two exact copies of the original DNA with slightly longer telomeres.
DNA Polymerase III is extremely accurate. Another DNA Polymerase acts as a proof reader, DNA Polymerase II. Once replication is complete, DNA Polymerase II travels down the entire link of the parental DNA strands looking for mismatched base pairs. Once it detects a mismatch, DNA Polymerase II removes the D-nucleotide from the new DNA strand and replaces it with the D-nucleotide complementary to the parental strand.
However , even this proofreading step is not completely accurate, missing mismatched base pair occurs approximately one out every billion times. These mistakes lead to nucleotide-level mutations. Such mutations are the only way in which new alleles are generated, and are usually neutral or deleterious relative to fitness. Occasionally these mutations will create beneficial phenotypes allowing these organisms higher probability of surviving and reproducing. Such mutations tend to amplify in future generations due to natural selection.
The Nucleus and DNA
Understanding genetics and DNA (deoxyribonucleic acid) can be quite challenging and so here are some notes on the basic foundations of genetics – we will look at the nucleus and its functions.
The nucleus is generally the largest organelle in an animal cell and contains the majority of the genetic material in that cell. It is separated from the surrounding cell through the presence of an envelope.
This envelope also contains pores, allowing controlled movement (using ATP) of substances in and out of the nucleus. Within the nucleus, there is also the nucleolus, which is the location for rRNA assembly.
The nucleus has 3 key functions:
What is DNA and how is it stored?
DNA contains the genetic information of a cell. It is an extremely stable double-stranded helix. It consists of nucleotides, of which there are 4: adenine, guanine, cytosine and thymine. These nucleotides are made up of a phosphate group, a deoxyribose sugar and a nitrogenous base. The bases have complementary and specific base pairing – adenine or pairs with thymine and guanine only pairs with cytosine.
The bases pair together with phosphodiester bonds and this leads to the formation of a sugar-phosphate backbone on the outside of the DNA – adding to its stability.
Obviously, DNA is very long and so there has to be an efficient way of storing it within the nucleus. This is done as follows:
- DNA is first wrapped around histone proteins to form a nucleosome
- Nucleosomes then form coils called solenoids
- These solenoids gather to form chromatin
- Chromatin is then further organised to form chromosomes (the main structures carrying the DNA) – chromosomes are only visible for a short period during metaphase in the cell cycle.
There are generally 22 pairs of chromosomes plus a pair of sex chromosomes (i.e. 23 pairs in total, or 46 chromosomes).
What are genes?
Genes are the crucial ‘unit’ of genetic information. A gene is essentially a section of DNA that codes for a particular protein. As an individual has 2 copies of each chromosome, they also have two versions of a gene (these may be the same or different). These versions are referred to as alleles.
Therefore, if these alleles are the same it is referred to as homozygous otherwise if they are different, it is referred to as heterozygous.
Genes contain exons, expressing sequences, and introns, interrupting sequences. When a gene is being processed, the introns are removed through splicing as they are of no use.
Already, from this, we can understand how changes in genes can have dramatic consequences. A single mutation of a base in a gene may result in a different protein product and, therefore, may result in a disease process.
This occurs during the S phase of the cell cycle (the process a cell goes through, including replication and growth). Replication occurs at several points and involves DNA polymerases (which will attempt to correct any potential errors).
There are three main steps to DNA replication: initiation, elongation, and termination. In order to fit within a cell’s nucleus, DNA is packed into tightly coiled structures called chromatin, which loosens prior to replication, allowing the cell replication machinery to access the DNA strands.
For better understanding we divided These three main steps into another six steps:
- 1) Initiation: Preparatory step
- Step 1: Replication fork formation.
- Step 2: Primer Binding
- Step 3: Synthesis of Leading and Lagging Strands
- Step 4: Remove Primer and Gap Fill
- Step 5: Proofreading
- Step 6: End Of the Replication
Bell, S. P. & Stillman, B. Nucleotide dependent recognition of chromosomal origins of DNA replication by a multi-protein complex. Nature 357, 128–134 ( 1992).
Diffley, J. F. X. & Cocker, J. H. Protein-DNA interactions at a yeast replication origin. Nature 357, 169–172 (1992).
Diffley, J. F. X., Cocker, J. H., Dowell, S. J. & Rowley, A. Two steps in the assembly of complexes at yeast replication origins in vivo. Cell 78, 303– 316 (1994).
Santocanale, C. & Diffley, J. F. X. ORC- and Cdc6-dependent complexes at active and inactive chromosomal replication origins in Saccharomyces cerevisiae. EMBO J. 15, 6671–6679 (1996).
Aparicio, O. M., Weinstein, D. M. & Bell, S. P. Components and dynamics of DNA replication complexes in S. cerevisiae: redistribution of MCM complexes and Cdc45p during S phase. Cell 91, 59–69 (1997).
Tanaka, T., Knapp, D. & Nasmyth, K. Loading of an Mcm protein onto DNA-replication origins is regulated by Cdc6p and CDKs. Cell 90, 649–660 (1997).
Cocker, J. H., Piatti, S., Santocanale, C., Nasmyth, K. & Diffley, J. F. X. An essential role for the Cdc6 protein in forming the pre-replicative complexes of budding yeast. Nature 379, 180–182 ( 1996).
Piatti, S., Lengauer, C. & Nasmyth, K. Cdc6 is an unstable protein whose de novo synthesis in G1 is important for the onset of S phase and for preventing a “reductional” anaphase in the budding yeast Saccharomyces cerevisiae. EMBO J. 14, 3788–3799 ( 1995).
Drury, L. S., Perkins, G. & Diffley, J. F. X. The Cdc4/34/53 pathway targets Cdc6p for proteolysis in budding yeast. EMBO J. 16, 5966– 5976 (1997).
Kearsey, S. E. & Labib, K. MCM proteins: evolution, properties, and role in DNA replication. Biochim. Biophys. Acta 1398, 113–136 ( 1998).
Tye, B. K. Mcm proteins in DNA replication. Annu. Rev. Biochem. 68, 649–686 (1999).
Yan, H., Merchant, A. M. & Tye, B.-K. Cell cycle-regulated nuclear localisation of MCM2 and MCM3, which are required for the initiation of DNA synthesis at chromosomal replication origins in yeast. Genes Dev. 7, 2149–2160 (1993).
Donovan, S., Harwood, J., Drury, L. S. & Diffley, J. F. X. Cdc6-dependent loading of Mcm proteins onto pre-replicative chromatin in budding yeast. Proc. Natl Acad. Sci. USA 94, 5611 –5616 (1997).
Liang, C. & Stillman, B. Persistent initiation of DNA replication and chromatin-bound MCM proteins during the cell cycle in cdc6 mutants . Genes Dev. 11, 3375–3386 (1997).
Young, M. R. & Tye, B. K. Mcm2 and Mcm3 are constitutive nuclear proteins that exhibit distinct isoforms and bind chromatin during specific cell cycle stages of Saccharomyces cerevisiae. Mol. Biol. Cell 8, 1587–1601 ( 1997).
Weinreich, M., Liang, C. & Stillman, B. The Cdc6p nucleotide-binding motif is required for loading Mcm proteins onto chromatin. Proc. Natl Acad. Sci. USA 96, 441–446 (1999).
Donaldson, A. D. et al. CLB5-dependent activation of late replication origins in S. cerevisiae. Mol. Cell 2, 173– 183 (1998).
Dahmann, C., Diffley, J. F. X. & Nasmyth, K. A. S-phase-promoting cyclin-dependent kinases prevent re-replication by inhibiting the transition of origins to a pre-replicative state. Curr. Biol. 5, 1257– 1269 (1995).
Detweiler, C. S. & Li, J. J. Ectopic induction of Clb2 in early G1 phase is sufficient to block prereplicative complex formation in Saccharomyces cerevisiae. Proc. Natl Acad. Sci. USA 95, 2384–2389 (1998).
Broek, D., Bartlett, R., Crawford, K. & Nurse, P. Involvement of p34 cdc2 in establishing the dependency of S phase on mitosis. Nature 349, 388– 393 (1991).
Hayles, J., Fisher, D., Woollard, A. & Nurse, P. Temporal order of S phase and mitosis in fission yeast is determined by the state of the p34 cdc2 -mitotic B cyclin complex. Cell 78, 813–822 (1994).
Moreno, S. & Nurse, P. Regulation of progression through the G1 phase of the cell-cycle by the rum1 + gene. Nature 367, 236–242 ( 1994).
Labib, K., Moreno, S. & Nurse, P. Interaction of cdc2 and rum1 regulates Start and S-phase in fission yeast. J. Cell Sci. 108, 3285 –3294 (1995).
Itzhaki, J. E., Gilbert, C. S. & Porter, A. C. Construction by gene targeting in human cells of a ‘‘conditional’’ CDC2 mutant that rereplicates its DNA. Nature Genet. 15, 258– 265 (1997).
Nishitani, H. & Nurse, P. p65 cdc18 plays a major role controlling the initiation of DNA replication in fission yeast. Cell 83, 397–405 ( 1995).
Jallepalli, P. V., Brown, G. W., Muzi-Falconi, M., Tien, D. & Kelly, T. J. Regulation of the replication initiator protein p65 cdc18 by CDK phosphorylation. Genes Dev. 11, 2767–2779 ( 1997).
Lopez-Girona, A., Mondesert, O., Leatherwood, J. & Russell, P. Negative regulation of cdc18 DNA replication protein by cdc2. Mol. Biol. Cell 9, 63–73 (1998).
Sánchez, M. M., Calzada, J. A. & Bueno, A. Functionally homologous DNA replication genes in fission and budding yeast. J. Cell Sci. 112, 2381 –2390 (1999).
Hennessy, K. M., Clark, C. D. & Botstein, D. Subcellular localization of yeast CDC46 varies with the cell cycle. Genes Dev. 4, 2252– 2263 (1990).
Dalton, S. & Whitbread, L. Cell-cycle-regulated nuclear import and export of Cdc47, a protein essential for initiation of DNA-replication in budding yeast. Proc. Natl Acad. Sci. USA 92, 2514–2518 (1995).
Amon, A., Irniger, S. & Nasmyth, K. Closing the cell cycle circle in yeast: G2 cyclin proteolysis initiated at mitosis persists until the activation of G1 cyclins in the next cycle. Cell 77, 1037–1050 (1994).
Tyers, M. The cyclin-dependent kinase inhibitor p40 SIC1 imposes the requirement for Cln G1 cyclin function at Start. Proc. Natl Acad. Sci. USA 93, 7772–7776 ( 1996).
Verma, R., Feldman, R. M. & Deshaies, R. J. SIC1 is ubiquitinated in vitro by a pathway that requires CDC4, CDC34, and cyclin/CDK activities. Mol. Biol. Cell 8, 1427–1437 ( 1997).
Desdouets, C. et al. Evidence for a Cdc6p-independent mitotic resetting event involving DNA polymerase α. EMBO J. 17, 4139 –4146 (1998).
Surana, U. et al. Destruction of the CDC28/CLB mitotic kinase is not required for the metaphase to anaphase transition in budding yeast. EMBO J. 12, 1969–1978 ( 1993).
Piatti, S., Bohm, T., Cocker, J. H., Diffley, J. F. X. & Nasmyth, K. Activation of S-phase promoting CDKs in late G1 defines a ‘‘point of no return’’ after which Cdc6 synthesis cannot promote DNA replication in yeast. Genes Dev. 10, 1516–1531 (1996).
Amon, A., Tyers, M., Futcher, B. & Nasmyth, K. Mechanisms that help the yeast cell cycle clock tick: G2 cyclins transcriptionally activate G2 cyclins and repress G1 cyclins. Cell 74, 993–1007 (1993).
Dohmen, R. J., Wu, P. & Varshavsky, A. Heat-inducible degron: a method for constructing temperature-sensitive mutants. Science 263, 1273– 1276 (1994).
Zwerschke, W., Rottjakob, H.-W. & Küntzel, H. The Saccharomyces cerevisiae CDC6 gene is transcribed at late mitosis and encodes a ATP/GTPase controlling S phase initiation. J. Biol. Chem. 269, 23351– 23356 (1994).
McInerny, C. J., Partridge, J. F., Mikesell, G. E., Creemer, D. P. & Breeden, L. L. A novel Mcm1-dependent element in the SWI4, CLN3, CDC6, and CDC47 promoters activates M/G1-specific transcription . Genes Dev. 11, 1277–1288 (1999).
Blow, J. J. & Laskey, R. A. A role for the nuclear envelope in controlling DNA replication within the cell cycle. Nature 332, 546–548 (1988).
Thommes, P., Kubota, Y., Takisawa, H. & Blow, J. J. The RLF-M component of the replication licensing system forms complexes containing all six MCM/P1 polypeptides. EMBO J. 16, 3312– 3319 (1997).
Hua, X. H., Yan, H. & Newport, J. A role for Cdk2 kinase in negatively regulating DNA replication during S phase of the cell cycle. J. Cell. Biol. 137 , 183–192 (1997).
Sanders Williams, R., Shohet, R. V. & Stillman, B. A human protein related to yeast Cdc6p. Proc. Natl Acad. Sci. USA 94, 142– 147 (1997).
Saha, P. et al. Human CDC6/Cdc18 associates with Orc1 and cyclin-cdk and is selectively eliminated from the nucleus at the onset of S phase. Mol. Cell Biol. 18, 2758–2767 ( 1998).
Jiang, W., Wells, N. J. & Hunter, T. Multistep regulation of DNA regulation by Cdk phosphorylation of HsCdc6. Proc. Natl Acad. Sci. USA 96, 6193–6198 (1999).
Peterson, B. O., Lukas, J., Sorenson, C. S., Bartek, J. & Helin, K. Phosphorylation of mammalian CDC6 by Cyclin A/CDK2 regulates its subcellular localization. EMBO J. 18, 396–410 ( 1999).
Sikorski, R. S. & Hieter, P. A system of shuttle vectors and yeast host strains designed for efficient manipulation of DNA in Saccharomyces cerevisiae. Genetics 122, 19–27 (1989).
Watch the video: Πώς αντιγράφεται το DNA? Και βασικά. Τι είναι το DNA? (May 2022).