Oscillations in the activity of cyclin-dependent kinases (CDKs) promote progression through the eukaryotic cell cycle. This review examines how proteolysis regulates CDK activity—by degrading CDK activators or inhibitors—and also how proteolysis may directly trigger the transition from metaphase to anaphase. Proteolysis during the cell cycle is mediated by two distinct ubiquitin-conjugation pathways. One pathway, requiring CDC34, initiates DNA replication by degrading a CDK inhibitor. The second pathway, involving a large protein complex called the anaphase-promoting complex or cyclosome, initiates chromosome segregation and exit from mitosis by degrading anaphase inhibitors and mitotic cyclins. Proteolysis therefore drives cell cycle progression not only by regulating CDK activity, but by directly influencing chromosome and spindle dynamics.

https://science.sciencemag.org/content/sci/274/5293/1652.full.pdf

How proteolysis drives the cell cycle

Doublecortin is a microtubule-associated protein and is expressed widely by migrating neurons.

https://www.cell.com/cell/pdf/0092-8674(81)90015-5.pdf

Feedback control of mitosis in budding yeast.

The tetratricopeptide repeat (TPR) motif is a protein-protein interaction module found in multiple copies in a number of functionally different proteins that facilitates specific interactions with a partner protein(s). Three-dimensional structural data have shown that a TPR motif contains two antiparallel alpha-helices such that tandem arrays of TPR motifs generate a right-handed helical structure with an amphipathic channel that might accommodate the complementary region of a target protein. Most TPR-containing proteins are associated with multiprotein complexes, and there is extensive evidence indicating that TPR motifs are important to the functioning of chaperone, cell-cycle, transcription, and protein transport complexes. The TPR motif may represent an ancient protein-protein interaction module that has been recruited by different proteins and adapted for specific functions. BioEssays 1999;21:932-939.

The tetratricopeptide repeat: a structural motif mediating protein-protein interactions.

Cell division is arrested in many organisms in response to DNA damage. Examinations of the genetic basis for this response in the yeast Saccharomyces cerevisiae indicate that the RAD9 gene product is essential for arrest of cell division induced by DNA damage. Wild-type haploid cells irradiated with x-rays either arrest or delay cell division in the G2 phase of the cell cycle. Irradiated G1 and M phase haploid cells arrest irreversibly in G2 and die, whereas irradiated G2 phase haploid cells delay in G2 for a time proportional to the extent of damage before resuming cell division. In contrast, irradiated rad9 cells in any phase of the cycle do not delay cell division in G2, but continue to divide for several generations and die. However, efficient DNA repair can occur in irradiated rad9 cells if irradiated cells are blocked for several hours in G2 by treatment with a microtubule poison. The RAD9-dependent response detects potentially lethal DNA damage and causes arrest of cells in G2 until such damage is repaired.

https://science.sciencemag.org/content/sci/241/4863/317.full.pdf

The RAD9 Gene Controls the Cell Cycle Response to DNA Damage in Saccharomyces cerevisiae

During a study of the genetics of nuclear migration in the filamentous fungus Aspergillus nidulans, we cloned a gene, nudF, which is required for nuclear migration during vegetative growth as well as development. The NUDF protein level is controlled by another protein NUDC, and extra copies of the nudF gene can suppress the nudC3 mutation. nudF encodes a protein with 42% sequence identity to the human LIS-1 (Miller-Dieker lissencephaly-1) gene, which is required for proper neuronal migration during brain development. This strong similarity suggests that the LIS-1 gene product may have a function similar to that of NUDF and supports previous findings to suggest that nuclear migration may play a role in neuronal migration.

/pdf/nudf-a-nuclear-migration-gene-in-aspergillus-nidulans-is-4cyxp8sza0.pdf

NudF, a nuclear migration gene in Aspergillus nidulans, is similar to the human LIS-1 gene required for neuronal migration.

Nuclear migration plays an important role in the growth and development of many organisms including the multinuclear fungus Aspergillus nidulans. We have identified four genes, nudA, nudC, nudF, and nudG, in which temperature-sensitive mutations affect nuclear distribution. In this report, we describe the cloning of the nudA gene by complementation of the mutant phenotype by using a chromosome VIII-specific cosmid library. A genomic fragment of nudA hybridized to an mRNA of approximately 14 kb. Sequencing analysis of nudA revealed four ATP-binding sites that are characteristic of the cytoplasmic dynein heavy chain. The amino acid sequence of the nudA gene product shows 52% overall identity with the rat brain cytoplasmic dynein heavy chain. Our study provides in vivo evidence that dynein, a microtubule motor molecule, plays a role in the nuclear migration process.

https://www.pnas.org/content/pnas/91/6/2100.full.pdf

Cytoplasmic dynein is involved in nuclear migration in Aspergillus nidulans

Forty-five temperature-sensitive mutants of Aspergillus nidulans which are defective in nuclear division, septation or distribution of nuclei along the mycelium have been isolated, and most have been subjected to complementation analysis and mapped to chromosome. Thirty-five of the mutants were unable to complete nuclear division at the restrictive temperature. Twenty-six of these mutants exhibited a co-ordinate drop in both spindle and chromosome mitotic indices at 42 °C, indicating that they fail to enter mitosis. These mutants have been assigned to the gene symbol nim. Nine mutants exhibited a co-ordinate rise in spindle and chromosome mitotic indices at 42 °C, indicating that they are arrested in mitosis. These mutants were assigned the gene symbol bim. Five mutants failed to form septa and were given the gene symbol sep; and five mutants had an abnormal nuclear distribution and were given the gene symbol nud. All of the mutations were recessive. Most of the mutants were in different complementation groups. Mutants in the same complementation groups were phenotypically similar, but phenotypically similar mutants were not necessarily or usually in the same complementation group. There was no evidence for genetic clustering of phenotypically similar mutants. The mutants were located on all eight chromosomes.

Mitotic mutants of Aspergillus nidulans.

The temperature-sensitive cell cycle mutation nimA5 causes nuclei of Aspergillus nidulans to be blocked in late G2 at restrictive temperature. Under these conditions the spindle pole body divides but does not separate and the mitotic index drops to zero. If nimA5 is blocked for more than one doubling time and then shifted from restrictive to permissive temperature, nuclei immediately enter mitosis, the mitotic spindle forms, and the chromosomes condense (Oakley, B. R., and N. R. Morris, 1983, J. Cell Biol., 96:1155-8). We have cloned the wild-type nimA gene by DNA-mediated complementation of the nimA5 mutant phenotype and have characterized nimA mRNA expression by Northern blot analysis. The transcript is 3.6 kb in length and is under tight nuclear cycle regulation. In synchronously dividing cells, the levels of nimA mRNA become elevated as cells enter mitosis and drop sharply as cells progress through mitosis. Cells blocked in S-phase with hydroxyurea have very low levels of nimA mRNA. Cells blocked in mitosis, either by the antimitotic agent benomyl or by the cell cycle mutation bimE7, maintain elevated levels of the nimA transcript. These data demonstrate not only that nimA is required for entry into mitosis, but because the transcript is normally expressed cyclically and is under tight cell cycle control, they suggest that nimA may play a regulatory role in the initiation of mitosis.

/pdf/regulation-of-the-mrna-levels-of-nima-a-gene-required-for-7a0y6tml2c.pdf

Regulation of the mRNA levels of nimA, a gene required for the G2-M transition in Aspergillus nidulans

To investigate the relationship between structure and function of kinesin-like proteins, we have identified by polymerase chain reaction (PCR) a new kinesin-like protein in the filamentous fungus Aspergillus nidulans, which we have designated KLPA. DNA sequence analysis showed that the predicted KLPA protein contains a COOH terminal kinesin-like motor domain. Despite the structural similarity of KLPA to the KAR3 and NCD kinesin-like proteins of Saccharomyces cerevisiae and Drosophila melanogaster, which also posses COOH-terminal kinesin-like motor domains, there are no significant sequence similarities between the nonmotor or tail portions of these proteins. Nevertheless, expression studies in S. cerevisiae showed that klpA can complement a null mutation in KAR3, indicating that primary amino acid sequence conservation between the tail domains of kinesin-like proteins is not necessarily required for conserved function. Chromosomal deletion of the klpA gene exerted no observable mutant phenotype, suggesting that in A. nidulans there are likely to be other proteins functionally redundant with KLPA. Interestingly, the temperature sensitive phenotype of a mutation in another gene, bimC, which encodes a kinesin-like protein involved in mitotic spindle function in A. nidulans, was suppressed by deletion of klpA. We hypothesize that the loss of KLPA function redresses unbalanced forces within the spindle caused by mutation in bimC, and that the KLPA and BIMC kinesin-like proteins may play opposing roles in spindle function.

/pdf/suppression-of-the-bimc4-mitotic-spindle-defect-by-deletion-2ys06w37o5.pdf

N R Morris

Papers

NudF, a nuclear migration gene in Aspergillus nidulans, is similar to the human LIS-1 gene required for neuronal migration.

Cytoplasmic dynein is involved in nuclear migration in Aspergillus nidulans

Mitotic mutants of Aspergillus nidulans.

Regulation of the mRNA levels of nimA, a gene required for the G2-M transition in Aspergillus nidulans

Suppression of the bimC4 mitotic spindle defect by deletion of klpA, a gene encoding a KAR3-related kinesin-like protein in Aspergillus nidulans