fizzy-related: Biological Overview | Evolutionary Homologs | Regulation | Developmental Biology | Effects of Mutation | References
Gene name - fizzy-related

Synonyms - retina aberrant in pattern

Cytological map position -

Function - signaling

Keywords - protein degradation, cell cycle, Anaphase-promoting complex/cyclosome (APC)

Symbol Symbol - fzr

FlyBase ID: FBgn0262699

Genetic map position - 1-

Classification - Trp-Asp repeat (WD-repeat) protein

Cellular location - presumably cytoplasmic



NCBI links: Precomputed BLAST | Entrez Gene | UniGene
Recent literature
Meghini, F., Martins, T., Tait, X., Fujimitsu, K., Yamano, H., Glover, D. M. and Kimata, Y. (2016). Targeting of Fzr/Cdh1 for timely activation of the APC/C at the centrosome during mitotic exit. Nat Commun 7: 12607. PubMed ID: 27558644
Summary:
A multi-subunit ubiquitin ligase, the anaphase-promoting complex/cyclosome (APC/C), regulates critical cellular processes including the cell cycle. To accomplish its diverse functions, APC/C activity must be precisely regulated in time and space. The interphase APC/C activator Fizzy-related (Fzr or Cdh1) is localized at centrosomes in animal cells. However, neither the mechanism of its localization nor its importance is clear. This study identified the centrosome component Spd2 as a major partner of Fzr in Drosophila. The localization of Fzr to the centriole during interphase depends on direct interaction with Spd2. By generating Spd2 mutants unable to bind Fzr, it was shown that centrosomal localization of Fzr is essential for optimal APC/C activation towards its centrosomal substrate Aurora A. Finally, it was shown that Spd2 is also a novel APC/C(Fzr) substrate. This study is the first to demonstrate the critical importance of distinct subcellular pools of APC/C activators in the spatiotemporal control of APC/C activity.
BIOLOGICAL OVERVIEW

The anaphase-promoting complex/cyclosome (APC) is a multisubunit ubiquitin ligase that targets several mitotic regulators for degradation and thereby triggers an exit from mitosis. APC activity is restricted to mitotic stages M and G1. This restriction is achieved by the cell cycle-dependent association of proteins encoded by fizzy and fizzy-related genes functioning during M and G1 respectively. fzr bears the accepted FlyBase designation of retina aberrant in pattern, because of a study showing that mutants show a disruption in eye patterning (Karpilow, 1989). Nevertheless, the alternative name fizzy-related is widely used and will be used in this essay. fzr, a conserved eukaryotic gene, negatively regulates the levels of cyclins A, B, and B3. These mitotic cyclins that bind and activate cdk1(cdc2) are rapidly degraded during exit from M and during G1. fzr is required for cyclin removal during G1 when the embryonic epidermal cell proliferation stops and during G2 during the cell cycle preceding salivary gland endoreduplication. Loss of fzr causes progression through an extra division cycle in the epidermis and inhibition of endoreduplication in the salivary gland, in addition to failure of cyclin removal. Conversely, premature fzr overexpression down-regulates mitotic cyclins, inhibits mitosis, and transforms mitotic cycles into endoreduplication cycles (Sigrist, 1997).

Binding of Cdc20 (the homolog of Fizzy in yeast and vertebrates) and Cdh1 (the homolog of Fizzy-related in yeast and vertebrates) to the APC is differentially regulated. APC-Cdc20 activity is present during mitosis and initiates the metaphase-anaphase transition. The association of Cdc20 with the APC requires phosphorylation of at least one subunit of the APC. Several mitotic kinases have been implicated in this phosphorylation. The dependency of APC phosphorylation on Cdc20 binding ensures that APC-Cdc20 is only active during mitosis. During prophase and prometaphase, APC-Cdc20 activity is furthermore restrained by the spindle checkpoint. This system monitors the presence of unattached kinetochores. Until kinetochores are bound by spindles, they serve as an assembly point for active Mad2 protein. Mad2 binds to Cdc20 and inhibits APC activity. Once all kinetochores are attached and chromosomes are aligned on the metaphase plate, Mad2 inhibition of APC-Cdc20 activity is released (Grosskortenhaus, 2002 and references therein).

Cdh1 is found in association with the APC during later stages of mitosis and G1. This interaction depends on the phosphorylation status of Cdh1 (Kramer, 2000; Zachariae, 1998). Only unphosphorylated Cdh1 is able to bind to and activate the APC (Kotani, 1999; Kramer, 2000). Cdk1 and Cdk2 mediate Cdh1 phosphorylation. Thus, only during stages of low Cdk kinase activity will Cdh1 activate the APC. These requirements are fulfilled during later stages of mitosis, when APC-Cdc20 has induced the degradation of mitotic cyclins, and during G1, when Cdk kinase activity is low. However, the G2 stage is also characterized by low Cdk kinase activity. How Cdh1-dependent APC activity is prevented in these situations has not been addressed so far (Grosskortenhaus, 2002 and references therein).

The mitotic cyclins in Drosophila (Cyclin A [CycA], Cyclin B [CycB], and Cyclin B3) are stable in interphase, degraded during mitosis, and continue to be unstable throughout G1. Cdc20Fzy is required for mitotic cyclin destruction at the metaphase-anaphase transition and is thought to mediate the bulk of cyclin degradation in the first 16 cell cycles in Drosophila. Mutants in fzy arrest in metaphase of cell cycle 16 when the maternal supply of Cdc20Fzy is exhausted. Overexpression of fzy does not cause abnormal cyclin destruction. Thus, Cdc20Fzy is not able to activate the APC at other cell cycle stages. This likely reflects the inability of Cdc20 to interact with unphosphorylated APC (Grosskortenhaus, 2002 and references therein).

Analysis in Drosophila demonstrates that fzr is expressed and of crucial importance when cells terminate cell proliferation during embryogenesis. Loss of fzr results in progression through an extra cell cycle in epidermal cells and in inhibition of endoreduplication in salivary glands. These deviations from the normal developmental cell cycle program are accompanied by a failure to down-regulate mitotic cyclins (cyclins A, B, and B3) that bind and activate cdk1(cdc2) kinase. Premature fzr overexpression, when epidermal cells still proliferate, down-regulates mitotic cyclins followed by inhibition of mitosis. All of these findings are consistent with the idea that Fzr activates degradation of mitotic cyclins and thereby prevents ectopic cdk1(cdc2) activity when cells become postmitotic (Sigrist, 1997).

The consequences of loss of fzr were examined with a deficiency deleting other genes in addition to fzr. However, in these deficient embryos, cell cycle defects occurred exclusively at stages and in tissues corresponding to the dynamic developmental program of fzr expression. Moreover, the various cell cycle defects were all corrected by expressing Hs-fzr in the deficient embryos at levels below the endogenous fzr expression as judged by in situ hybridization. Finally, except for correcting the cell cycle defects in tissues that normally express fzr, this moderate Hs-fzr expression has no effects on cell cycle progression in deficient embryos (Sigrist, 1997).

Ectopic cyclin E expression as well as failure to express the cyclin E/cdk2 inhibitor p27DAP in the embryonic epidermis results in ectopic accumulation of mitotic cyclins and in progression through an ectopic division cycle instead of a G1 arrest. Therefore, ectopic cyclin E/cdk2 activity in the postmitotic epidermal cells has the same phenotypic consequences as loss of fzr, suggesting that Fzr might act primarily as a negative regulator of cyclin E/cdk2 and indirectly on mitotic cyclins. All of the following findings, however, argue strongly against this interpretation. By immunolabeling with anti-cyclin E and anti-DAP antibodies, altered expression in either fzr-deficient embryos or after prd-Gal4-directed UAS-fzr expression was not detected (Sigrist, 1997).

In addition, while ectopic UAS-dap expression in the salivary gland during the larval stages inhibits endoreduplication effectively, UAS-fzr expression has no effect. Finally, when UAS-fzr expression is directed to imaginal disc cells during the larval stages, endoreduplication instead of the normal mitotic proliferation results, as also observed in CycA and Cdc2 mutants, but not in CycE mutants (Sigrist, 1997).

The onset of the developmentally programmed physiological endoreduplication in the salivary gland, therefore, might depend on the inactivation of cdk1 complexes. This inactivation might be achieved by the down-regulation of mitotic cyclins caused by the especially high levels of fzr expression that are observed transiently in the salivary gland before the onset of the first endoreduplication S phase. In the fzr-deficient embryos, mitotic cyclins continue to accumulate and cdk1 activity is expected to be maintained, resulting in the inhibition of endoreduplication (Sigrist, 1997).

While the inactivation of cdk1 is thought to establish the competence to initiate another round of DNA replication, the actual onset of DNA replication is dependent on the activation of cdk1 in yeast and cdk2 in higher eukaryotes. Particular cyclins, Clb5 and Clb6 in yeast and cyclins E and A in higher eukaryotes, are normally involved in this activation under physiological conditions. However, DNA replication can be activated in yeast and higher eukaryotes by cyclin/cdk1 complexes that play little or no role during the G1/S transition under physiological conditions. Moreover, ectopic expression of Drosophila cyclin A, which appears to bind only to cdk1 and not to cdk2, is definitively capable of driving cells from G1 into S. Premature activation of cdk1 must presumably be prevented when cells have to be maintained in the G1 phase. The entry into an additional S phase that is observed in the epidermis of fzr-deficient embryos, therefore, might also result from the failure to down-regulate the mitotic cyclins (Sigrist, 1997).

fzr was identified because of its similarity to fzy, which is required for progression beyond metaphase and mitotic cyclin degradation. fzr and fzy encode highly similar proteins with seven WD repeats in the C-terminal region. WD repeats are found in many proteins with diverse biological function. They are also found in budding yeast Cdc4p, which is required for the ubiquitin-dependent proteolysis of important cell cycle regulators. CDC4 acts in a pathway with CDC53, and interestingly, mutations in cul-1, a C. elegans homolog of CDC53, have recently been shown to result in a similar inability to arrest cell proliferation at the appropriate developmental stage as also apparent in the fzr-deficient embryos (Sigrist, 1997).

The closest yeast relative of fzr, however, is not CDC4 but HCT1, which is required for proteolysis of Clb2p, a budding yeast B-type cyclin with a characteristic destruction box as also present in A- and B-type cyclins of higher eukaryotes. Drosophila fzr appears to be unable to provide HCT1 function in yeast. It remains to be demonstrated that Fzr activates the proteolytic degradation of mitotic cyclins. However, the abnormal cyclin accumulation in both fzr and fzy mutants is not associated with an apparent increase in transcript levels, indicating that Fzr and Fzy down-regulate mitotic cyclins at a posttranscriptional level. If Fzy and Fzr trigger cyclin proteolysis as suspected, it will be important to clarify their functional relationship with proteins known to be required for the ubiquitin-dependent degradation of mitotic cyclins, with Cse1p and the anaphase-promoting complex (APC), a complex composed of several proteins. In addition, it will be interesting to evaluate the relationship of fzr and roughex, an unrelated Drosophila gene with a similar function required at other developmental stages (Sigrist, 1997).

Double mutant analyses demonstrate that Fzy and Fzr are specialized for the down-regulation of mitotic cyclins during either M phase or interphase, respectively. fzy is expressed in proliferating cells and is required for progression beyond metaphase and mitotic cyclin degradation; fzr transcripts accumulate when cells become postmitotic and are required for cyclin down-regulation in G1 during cell cycle exit and in G2 before endoreduplication but not during mitosis. It will be interesting to learn whether fzr is required in proliferating cells progressing through cell cycles with G1 phases (which do not occur during Drosophila embryogenesis). Since Fzy and Fzr promote down-regulation of mitotic cyclins in different cell cycle phases, they might have evolved to respond to different regulatory inputs. A fraction of Fzy is modified precisely during the metaphase/anaphase transition when mitotic cyclin degradation starts. Cyclin degradation during mitosis, and perhaps Fzy activity as well, is controlled by checkpoint mechanisms that monitor spindle integrity and chromosome attachment (Sigrist, 1997)

In contrast, degradation of mitotic cyclins during interphase appears to be important for cell cycle exit and entry into endoreduplication. fzr transcription is regulated by developmental cues that stop embryonic cell proliferation. In addition, Fzr appears to be regulated as well. Observations suggest that Fzr is negatively regulated by cyclin E/cdk2 activity, which in turn is also controlled by the developmental cues that stop embryonic cell proliferation. These developmental cues result in CycE down-regulation and dacapo up-regulation at the stage where cell proliferation is to be arrested. Therefore, the resulting inhibition of cyclin E/cdk2 activity might activate Fzr and thereby prevent mitotic cyclin accumulation and ectopic cdk1 activity when cells become postmitotic (Sigrist, 1997).

It is likely that FZY and Fzr are involved in the degradation of proteins other than the known A- and B-type cyclins. It is thought that the degradation of proteins like the fission yeast cut2 protein is required for sister chromatid separation in mitosis, and fzy is clearly required not only for cyclin degradation but also for sister chromatid separation in mitosis. It is possible, therefore, that Fzy and Fzr trigger the proteolytic degradation of different subsets of proteins (Sigrist, 1997).

In summary, the conserved fzr gene might be generally important in higher eukaryotes for transitions in the developmental cell cycle program (Sigrist, 1997).


GENE STRUCTURE

cDNA clone length - 2605

Bases in 5' UTR - 467

Exons - 5

Bases in 3' UTR - 701


PROTEIN STRUCTURE

Amino Acids - 478

Structural Domains

Drosophila fizzy is required for cell cycle progression beyond metaphase and for mitotic degradation of A- and B-type cyclins (Dawson, 1993; Dawson, 1995; Sigrist, 1995). The Xenopus fzr gene was identified in an attempt to identify a fzy homolog in Xenopus, an ideal organism for biochemical analyses of cyclin proteolysis. Subsequent analyses have revealed the presence of both, fzy and fzr, in Drosophila and Xenopus. Sequencing projects have identified apparent fzy and fzr homologs in S. pombe, C. elegans, and vertebrates (Sigrist, 1997).

The protein products encoded by the fzy and fzr cDNAs are highly similar in their C-terminal domains, which are composed of seven WD repeats. These repeats, which are found in many functionally diverse proteins, were first identified in ß-transducin. A crystallographic analysis has demonstrated that these WD- or ß-transducin repeats form a seven propeller structure that provides protein-protein interaction faces in trimeric G-protein complexes. In the N-terminal domain, similarities between fzy and fzr are restricted to a few motifs (Sigrist, 1997).


retina aberrant in pattern/fizzy-related: Evolutionary Homologs | Regulation | Developmental Biology | Effects of Mutation | References

date revised: 15 February 2002

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