Plant Cell Biology Lecture 4 Part II - Fall2024 PDF

040407221: PLANT CELL BIOLOGY ‫بيولوچيا الخلية النباتية‬ Lecture 4 Part II DR. FATMA M. ASHOUR 1 Dr.Fatma Ashour  CELL CYCLE CONTROL SYSTEMS The eukaryotic cell cycle is driven by cell cycle “engines”, a set of interacting p...

040407221: PLANT CELL BIOLOGY ‫بيولوچيا الخلية النباتية‬ Lecture 4 Part II DR. FATMA M. ASHOUR 1 Dr.Fatma Ashour  CELL CYCLE CONTROL SYSTEMS The eukaryotic cell cycle is driven by cell cycle “engines”, a set of interacting proteins, the cyclin- dependent kinases (Cdks). CDKs are relatively small proteins, with molecular weights ranging from 34 to 40 kDa, and contain little more than the kinase domain. By definition, a CDK binds a regulatory protein called a cyclin. Without cyclin, CDK has little kinase activity; only the cyclin-CDK complex is an active kinase. 1- Cyclin 2- Kinase (cyclin-dependent kinase) 1 3- Inhibitor (cyclin-dependent kinase inhibitor) 3 Dr.Fatma Ashour 2  CELL CYCLE CONTROL SYSTEMS A kinase is an enzyme that catalyzes the transfer of phosphate groups from high-energy, phosphate-donating molecules to specific substrates. This process is known as phosphorylation, where the substrate gains a phosphate group and the high-energy ATP molecule donates a phosphate group. Cyclin is a family of proteins that controls the progression of a cell through the cell cycle by activating cyclin-dependent kinase (CDK) enzymes or group of enzymes required for synthesis of cell cycle. Dr.Fatma Ashour 3  CELL CYCLE CONTROL SYSTEMS Other proteins act as rate-limiting steps in cell cycle progression and are able to induce cell cycle arrest at defined stages (checkpoints). The cyclic pattern of cyclin expression to produce progression through the cell cycle is under transcriptional control. Dr.Fatma Ashour 4  figure 1. Cell cycle and checkpoints Dr.Fatma Ashour 5 FIGURE 2 : CELL CYCLE CONTROL SYSTEMS Dr.Fatma Ashour 6  Activation of CdKs requires their dephosphorylation; kinase activity can also be blocked by cyclin- dependent kinase inhibitors (CKIs). One CKI, known as P16, is in fact a tumor suppressor, deficiencies in which are associated with many cancers. In yeast, there is a single kinase that interacts with different cyclins to promote different cell- cycle transitions. In mammals and plants, the situation is more complicated, with different kinases binding different cyclins to promote the different transitions. Dr.Fatma Ashour 7 There are three major classes of cyclin- CDK complexes that control passage through the cell cycle: 1- G1, cyclin- CDK 2- S-phase, cyclin- CDK 3-Mitotic cyclin- CDK complexes. When cells are stimulated to replicate, G1 cyclin- CDK complexes are expressed first. Dr.Fatma Ashour 8  G1 CYCLIN-CDK COMPLEXES G1 cyclin-CDK complexes prepare the cell for the S phase by activating transcription factors. These factors promote transcription of: 1- Genes encoding enzymes required for DNA synthesis. 2- The genes encoding S-phase cyclins and CDKs. Note: The activity of S-phase cyclin-CDK complexes initially held in check by inhibitors. Dr.Fatma Ashour 9 FIGURE 3: Control of the G1 → S phase transition in S. cerevisiae by regulated proteolysis of the S-phase inhibitor Sic1. The S-phase cyclin-CDK complexes begin to accumulate in G1, but are inhibited by Sic1. This inhibition prevents initiation of DNA replication until the cell is fully prepared. G1 cyclin-CDK complexes assembled in late G1 phosphorylate Sic1 (step 1), marking it for polyubiquitination, and subsequent proteasomal degradation (step 2). The active S-phase cyclin-CDK complexes then trigger initiation of DNA synthesis (step 3) by phosphorylating substrates that remain to be identified. [Adapted from R. W. King et al., 1996, Science 274:1652. Dr.Fatma Ashour 10 In late G1, the G1 cyclin-CDK complexes induce degradation of the S-phase inhibitors releases active S-phase cyclin-CDK complexes. The function of active S-phase cyclin-CDK complexes : 1- activate initiation of DNA replication 2- prevent (inhibit) re-replication of the DNA molecule Because of this inhibition, each chromosome is replicated just once during passage through the cell cycle, ensuring that the proper chromosome number is maintained in the daughter cells. Dr.Fatma Ashour 11 The most important exception to this is Endoreduplication, in which successive rounds of replication occur without cell or chromosomal division This is the process by which polytene chromosomes are formed. Interband (euchromatin) Band (heterochromatin) The normal mitotic cell cycle controls are modified to that DNA replication is not followed by mitosis. Dr.Fatma Ashour 12 3- S cyclin- CDK activates helicases, which unwind the parental DNA strands. 4- A complex of DNA polymerase (pol. α ) and primase initiates the synthesis of daughter strands. 5- DNA polymerase δ plus its accessory factors, elongate daughter strands initiated by pol α - primase. Dr.Fatma Ashour 13 MITOTIC CYCLIN- CDK COMPLEXES Mitotic cyclin- CDK complexes are synthesized during the S phase and G2, but their activities are held until DNA synthesis is completed. Mitotic cyclin-CDK complexes is activated by dephosphorylation by phosphatases. Active M cyclin- CDK activates early mitotic events, these events are: 1- Chromosome condensation, 2- Retraction of the nuclear envelope, 3- Assembly of the mitotic spindle apparatus, and 4-Alignment of condensed chromosomes at the metaphase plate. Dr.Fatma Ashour 14 At metaphase, the spindle is in a state of tension with forces pulling the two kinetochores toward the opposite spindle poles balanced by forces pushing the spindle poles apart. Sister chromatids do not separate because they are held together at their centromere and multiple positions along the chromosome arms by multiprotein complexes called cohesions. Dr.Fatma Ashour 15 THE ANAPHASE PROMOTING COMPLEX (APC) This sequence of events initiates anaphase by freeing sister chromatids to segregate to opposite spindle poles. Late in anaphase, the APC also directs the degradation of the mitotic cyclins. The degradation of the mitotic cyclins by APC is inhibited until the segregating chromosomes have reached the proper location in the dividing cell. Degradation of the mitotic cyclins leads to decrease in mitotic CDK activity. Dr.Fatma Ashour 16 FIGURE 4: MODEL FOR CONTROL OF ENTRY INTO ANAPHASE BY APC- REGULATED DEGRADATION OF THE COHESIN LINK BETWEEN SISTER CHROMATIDS. Dr.Fatma Ashour 17  THE ANAPHASE PROMOTING COMPLEX (APC) As a result, the decrease in mitotic CDK activity permits to: 1- de- condense of separated chromosomes , 2- the nuclear envelope re-forms around daughter-cell nuclei, and 3- the Golgi apparatus reassembles. Finally during Telophase; the cytoplasm divides at cytokinesis, yielding the two daughter cells. Passage through three critical cell-cycle transitions: G1 → S phase, metaphase → anaphase, and anaphase → telophase and cytokinesis, is irreversible because these transitions are triggered by the regulated degradation of proteins, and irreversible process. As a consequence, cells are forced to traverse the cell cycle in one direction only. Dr.Fatma Ashour 18 MEIOSIS: A SPECIAL TYPE OF CELL DIVISION Meiosis involves one cycle of chromosome replication followed by two cycles of cell division to produce haploid germ cells from a diploid premeiotic cell. During meiosis I, replicated homologous chromosomes pair along their lengths in a process called synapsis. At least one recombination between chromatids of homologous chromosomes almost in variably occurs. Most of the cell-cycle proteins that function in mitotically dividing cells also function in cells undergoing meiosis, but some proteins are unique to meiosis. Dr.Fatma Ashour 19 Figure 5. Dr.Fatma Ashour 20  MEIOSIS: A SPECIAL TYPE OF CELL DIVISION In S. cerevisiae, expression of G1 cyclins is repressed throughout meiosis. Meiosis-specific Ime2 performs the function of G1 cyclin-CDK complexes in promoting initiation of DNA replication during meiosis I. DNA replication does not occur during meiosis II because neither Ime2 nor G1 cyclins are expressed. In S. cerevisiae, recombination (crossing over) between chromatids of homologous parental chromatids and cohesin cross-links between chromatids distal to the crossover are responsible for synapsis of homologous chromosomes during prophase and metaphase of meiosis I. A specialized cohesin subunit, Rec8, replaces the Scc1 cohesin subunit during meiosis. Dr.Fatma Ashour 21 MEIOSIS: A SPECIAL TYPE OF CELL DIVISION During early anaphase of meiosis I, Rec8 in the chromosome arms is cleaved, but a meiosis-specific protein associated with the kinetochore protects Rec8 in the region of the centromere from cleavage. As a result, the chromatids of homologous chromosomes remain associated during segregation in meiosis I. Cleavage of centromeric Rec8 during anaphase of meiosis II allows individual chromatids to segregate into germ cells. Monopolin, another meiosis-specific protein, is required for both chromatids of homologous chromosomes to associate with microtubules emanating from the same spindle poles during meiosis I. Dr.Fatma Ashour 22 FIGURE 6. Cohesin function during mitosis and meiosis. (a) During mitosis, sister chromatids generated by DNA replication in the S phase are initially associated by cohesin complexes along the full length of the chromatids. During chromosome condensation, cohesin complexes (yellow) become restricted to the region of the centromere at metaphase, as depicted here. Once separase cleaves the Scc1 cohesin subunit, sister chromatids can separate, marking the onset of anaphase. (b) In metaphase of meiosis I, crossing over between maternal and paternal chromatids produces synapsis of homologous parental chromosomes. The chromatids of each replicated chromosome are cross-linked by cohesin complexes along their full length. Rec8, a meiosis- specific homolog of Scc1, is cleaved in chromosome arms but not in the centromere, allowing homologous chromosome pairs to segregate to daughter cells. Centromeric Rec8 is cleaved during meiosis II, allowing individual chromatids to segregate to daughter cells. [Modified from F. Uhlmann, 2001, Curr. Opin. Cell Biol. 13:754.] Dr.Fatma Ashour 23  MEIOTIC ARREST In the mitotic cell cycle, the main variable in the length of the cell cycle is the length of G1, and once the cell is committed to divided, it proceeds through S, G2 and mitosis without delay. Similarly, male meiosis is generally a continuous process, designed to produce vast quantities of spermatocytes with the minimum delay. Female germ cells, however, can arrest in meiosis, often for years, at a variety of different stages, according to the species. - In many invertebrates this arrest is at metaphase I. - In most vertebrates it is at metaphase II. The stimulus for completion of meiosis is usually fertilization. Dr.Fatma Ashour 24  MEIOTIC ARREST Meiotic arrest also occurs commonly at diplotene. In mammals, female meiosis starts in the embryo, and proceeds as far as diplotene, when the chromosomes become diffuse and the cells are referred to as being in the dictyate stage. This arrest is under hormonal control, and the oocyte recommences growth and passage through meiosis in response to luteinizing hormone in adult life. The dictyate or dictyotene is a prolonged resting phase in oogenesis Dr.Fatma Ashour 25