Casein kinase II phosphorylates Ser<sup>468</sup> in the PEST domain of the <i>Drosophila</i> IκB homologue cactus

Packman, Leonard C.; Kubota, Ken; Parker, J. S. F.

doi:10.1016/s0014-5793(96)01324-5

Cited by 35 publications

(30 citation statements)

References 23 publications

Supporting

Mentioning

Contrasting

Unclassified

Order By: Relevance

“…Only two of the three PEST domain serines reside in a consensus CKII phosphorylation site. Furthermore, although we find that mutation of any single PEST domain serine does not eliminate CKII-mediated phosphorylation of Cactus (data not shown), Gay and colleagues have recently found that Ser-468 is the predominant site of modification in vitro (Packman et al 1997). We expect, therefore, that CKII phosphorylation introduces less negative charge than three serine-to-glutamate substitutions.…”

Section: Wild-type Activity Of the Cactus Pest Domain In Embryos Requcontrasting

confidence: 60%

A role for CKII phosphorylation of the Cactus PEST domain in dorsoventral patterning of the Drosophila embryo

Liu¹,

Galindo²,

Wasserman³

1997

Genes Dev.

View full text Add to dashboard Cite

Regulated proteolysis of Cactus, the cytoplasmic inhibitor of the Rel-related transcription factor Dorsal, is an essential step in patterning of the Drosophila embryo. Signal-induced Cactus degradation frees Dorsal for nuclear translocation on the ventral and lateral sides of the embryo, establishing zones of gene expression along the dorsoventral axis. Cactus stability is regulated by amino-terminal serine residues necessary for signal responsiveness, as well as by a carboxy-terminal PEST domain. We have identified Drosophila casein kinase II (CKII) as a Cactus kinase and shown that CKII specifically phosphorylates a set of serine residues within the Cactus PEST domain. These serines are phosphorylated in vivo and are required for wild-type Cactus activity. Conversion of these serines to alanine or glutamic acid residues differentially affects the levels and activity of Cactus in embryos, but does not inhibit the binding of Cactus to Dorsal. Taken together, these data indicate that wild-type axis formation requires CKII-catalyzed phosphorylation of the Cactus PEST domain. A nuclear concentration gradient of the transcription factor Dorsal defines the dorsoventral axis of the Drosophila embryo (for review, see Chasan and Anderson 1993). Dorsal is a member of the Rel family of proteins that includes mammalian NF-B, the avian oncogene vrel, and the cellular proto-oncogene c-rel (Steward 1987; for review, see Verma et al. 1995). Transcriptional activity of Rel proteins is controlled at the level of subcellular localization. More specifically, ankyrin-repeat containing inhibitory proteins such as Cactus, IB␣, and IB␤ retain Rel proteins in the cytoplasm by direct interaction. On stimulation, signal transduction targets the inhibitors for degradation, freeing the Rel-proteins for translocation into nuclei.A maternally encoded signal transduction pathway directs the asymmetric nuclear translocation of Dorsal in the embryo (for review, see Chasan and Anderson 1993). A protease cascade active in the ventral portion of the extraembryonic space cleaves, and thereby activates, the Spä tzle protein. Spä tzle then acts as an extracellular ligand, binding the transmembrane receptor Toll. This asymmetric activation of Toll initiates intracellular signal transduction via the protein kinase Pelle that drives Cactus degradation, thereby freeing Dorsal for nuclear import. On translocation into ventral and lateral nuclei, Dorsal establishes embryonic polarity by activating transcription of ventral-specific genes while repressing dorsal-specific loci.The pathway governing subcellular localization of Drosophila Dorsal displays striking similarities to at least one pathway that controls the partitioning of mammalian NF-B between the nucleus and cytoplasm (Wasserman 1993;Cao et al. 1996). At least four of the proteins in the Drosophila dorsoventral pathway-Toll, Pelle, Dorsal, and Cactus-have structural and functional counterparts in the response of lymphocytes and other cell types to the cytokine interleukin-1 (IL-1)-the IL-1 recepto...

show abstract

Section: Wild-type Activity Of the Cactus Pest Domain In Embryos Requcontrasting

confidence: 60%

A role for CKII phosphorylation of the Cactus PEST domain in dorsoventral patterning of the Drosophila embryo

Liu¹,

Galindo²,

Wasserman³

1997

Genes Dev.

View full text Add to dashboard Cite

show abstract

“…This sequence, hereafter referred to as the SPXSS-SDXE motif is located within a region with a high PEST score (37). PEST-rich sequences behave as cisacting signals that regulate protein turnover and have been suggested to be activated via phosphorylation (27,31). The Drosophila m5, m7, and m8 proteins were shown to associate with and be phosphorylated by protein kinase CK2 at their conserved SPXSS-SDXE sequences.…”

Section: Resultsmentioning

confidence: 99%

“…Regardless of the exact nature of the events induced by MG132, the observation that Hes6 is prone to proteolytic degradation is in agreement with the presence of an evolutionarily conserved PEST domain containing an SPXSS-SDXE subdomain that includes a resident consensus protein kinase CK2 phosphorylation site at Ser183. The presence of PEST domains is characteristic of proteins that undergo increased turnover, and phosphorylation of PEST sequences by protein kinase CK2 was shown to negatively affect intrinsic protein stability (22,27,31). The Drosophila Hes family members Enhancer of split m5, m7, and m8 share with Hes6 a similar SPXSS-SDXE motif within a carboxy-terminal region characterized by a high PEST score.…”

Section: Vol 23 2003 Molecular Mechanisms Of Hes6 Function 6931mentioning

confidence: 99%

Hes6 Promotes Cortical Neurogenesis and Inhibits Hes1 Transcription Repression Activity by Multiple Mechanisms

Gratton

Torban

Jasmin

et al. 2003

Molecular and Cellular Biology

112

150

View full text Add to dashboard Cite

Hes1 is a mammalian basic helix-loop-helix transcriptional repressor that inhibits neuronal differentiation together with corepressors of the Groucho (Gro)/Transducin-like Enhancer of split (TLE) family. The interaction of Hes1 with Gro/TLE is mediated by a WRPW tetrapeptide present in all Hairy/Enhancer of split (Hes) family members. In contrast to Hes1, the related protein Hes6 promotes neuronal differentiation. Little is known about the molecular mechanisms that underlie the neurogenic activity of Hes6. It is shown here that Hes6 antagonizes Hes1 function by two mechanisms. Hes6 inhibits the interaction of Hes1 with its transcriptional corepressor Gro/TLE. Moreover, it promotes proteolytic degradation of Hes1. This effect is maximal when both Hes1 and Hes6 contain the WRPW motif and is reduced when Hes6 is mutated to eliminate a conserved site (Ser183) that can be phosphorylated by protein kinase CK2. Consistent with these findings, Hes6 inhibits Hes1-mediated transcriptional repression in cortical neural progenitor cells and promotes the differentiation of cortical neurons, a process that is normally inhibited by Hes1. Mutation of Ser183 impairs the neurogenic ability of Hes6. Taken together, these findings clarify the molecular events underlying the neurogenic function of Hes6 and suggest that this factor can antagonize Hes1 activity by multiple mechanisms.In the developing mammalian central nervous system (CNS), differentiated neuronal and glial cells derive from multipotent neural progenitor cells located in the proliferative zone of the neural tube. The commitment of these progenitor cells to the neuronal lineage is regulated by the antagonistic activities of a number of positively and negatively acting transcription factors containing the basic helix-loop-helix (bHLH) DNA-binding and dimerization motif (reviewed in references 2 and 18). Neurogenic bHLH factors include several evolutionarily conserved molecules related to the proneural proteins Atonal and Achaete-Scute of Drosophila (8, 13, 21). They function by forming heterodimers with the ubiquitous bHLH protein E47. These dimers bind to DNA sequences commonly referred to as E boxes (CANNTG) and transactivate the expression of genes that promote the acquisition of the neuronal fate (17, 32).Antineurogenic bHLH factors include members of the Hairy/Enhancer of split (Hes) family (1,26,32). In contrast to proneural proteins, Hes factors like Hes1 and Hes5 mediate transcriptional repression and bind preferentially to DNA sequences referred to as N boxes (CACNAG) (32). They are thought to inhibit neuronal differentiation by antagonizing the neurogenic activity of the proneural proteins via multiple mechanisms, including direct involvement in the negative regulation of proneural gene expression (4,20) and inhibition of the activity of E47-proneural protein heterodimers (1,3,32). Genetic perturbations that alter the normal balance of the activities of proneural and antineurogenic bHLH proteins have dramatic effects on CNS development in vivo, underscoring th...

show abstract

“…Dorsoventral patterning in the Drosophila embryo involves the activities of a transcription factor, Dorsal, and its inhibitor, Cactus (73). Upon receiving the inductive signal, Cactus appears to be phosphorylated by CKII within a motif with a high PEST score (74) and undergoes degradation, thus allowing Dorsal to translocate to the nucleus. A mechanistically similar situation appears to regulate the NF-B/C-Rel family of transcription factors in humans (75,76).…”

mentioning

confidence: 99%

Drosophila melanogaster Casein Kinase II Interacts with and Phosphorylates the Basic Helix-Loop-Helix Proteins m5, m7, and m8 Derived from the Enhancer of split Complex

Trott

Kalive

Paroush

et al. 2001

Journal of Biological Chemistry

View full text Add to dashboard Cite

Drosophila melanogaster casein kinase II (DmCKII) is composed of catalytic (␣) and regulatory (␤) subunits associated as an ␣ 2 ␤ 2 heterotetramer. Using the twohybrid system, we have screened a D. melanogaster embryo cDNA library for proteins that interact with Dm-CKII␣. One of the cDNAs isolated in this screen encodes m7, a basic helix-loop-helix (bHLH)-type transcription factor encoded by the Enhancer of split complex (E(spl)C), which regulates neurogenesis. m7 interacts with DmCKII␣ but not with DmCKII␤, suggesting that this interaction is specific for the catalytic subunit of Dm-CKII. In addition to m7, we demonstrate that DmCKII␣ also interacts with two other E(spl)C-derived bHLH proteins, m5 and m8, but not with other members, such as m3 and mC. Consistent with the specificity observed for the interaction of DmCKII␣ with these bHLH proteins, sequence alignment suggests that only m5, m7, and m8 contain a consensus site for phosphorylation by CKII within a subdomain unique to these three proteins. Accordingly, these three proteins are phosphorylated by DmCKII␣, as well as by the ␣ 2 ␤ 2 holoenzyme purified from Drosophila embryos. In line with the prediction of a single consensus site for CKII, replacement of Ser 159 of m8 with either Ala or Asp abolishes phosphorylation, identifying this residue as the site of phosphorylation. We also demonstrate that m8 forms a direct physical complex with purified DmCKII, corroborating the observed two-hybrid interaction between these proteins. Finally, substitution of Ser 159 of m8 with Ala attenuates interaction with DmCKII␣, whereas substitution with Asp abolishes the interaction. These studies constitute the first demonstration that DmCKII interacts with and phosphorylates m5, m7, and m8 and suggest a biochemical and/or structural basis for the functional equivalency of these bHLH proteins that is observed in the context of neurogenesis. Casein kinase II (CKII)1 is a ubiquitous protein kinase that is highly conserved among eukaryotes (1, 2) and is capable of functioning as an oncogene in mammals (3). CKII is composed of catalytic (␣) and regulatory (␤) subunits that combine to form an ␣ 2 ␤ 2 holoenzyme. With the exceptions of Drosophila melanogaster (4), Caenorhabditis elegans (5), and Schizosaccharomyces pombe (6), CKII from most eukaryotic organisms contains two ␣ subunits, ␣ and ␣Ј, that are encoded by distinct genes. In contrast, ␤ subunit heterogeneity has been documented via protein microchemical approaches in Saccharomyces cerevisiae (7) and via molecular/genetic approaches in Arabidopsis thaliana (8) and D. melanogaster (9).CKII preferentially phosphorylates Ser/Thr residues in an hyperacidic context (10), although phosphorylation of Tyr has been documented in at least one case, i.e. yeast Fpr3 (11). Analysis of the phosphorylation of synthetic peptides suggests that the consensus site for phosphorylation by CKII can best be described as (S/T)(D/E)X(D/E) (10). Consistent with this, a number of proteins critical for transcription, cell cycle regulation, and s...

show abstract

Casein kinase II phosphorylates Ser⁴⁶⁸ in the PEST domain of the Drosophila IκB homologue cactus

Cited by 35 publications

References 23 publications

A role for CKII phosphorylation of the Cactus PEST domain in dorsoventral patterning of the Drosophila embryo

A role for CKII phosphorylation of the Cactus PEST domain in dorsoventral patterning of the Drosophila embryo

Hes6 Promotes Cortical Neurogenesis and Inhibits Hes1 Transcription Repression Activity by Multiple Mechanisms

Drosophila melanogaster Casein Kinase II Interacts with and Phosphorylates the Basic Helix-Loop-Helix Proteins m5, m7, and m8 Derived from the Enhancer of split Complex

Contact Info

Product

Resources

About

Casein kinase II phosphorylates Ser468 in the PEST domain of the Drosophila IκB homologue cactus

Cited by 35 publications

References 23 publications

A role for CKII phosphorylation of the Cactus PEST domain in dorsoventral patterning of the Drosophila embryo

A role for CKII phosphorylation of the Cactus PEST domain in dorsoventral patterning of the Drosophila embryo

Hes6 Promotes Cortical Neurogenesis and Inhibits Hes1 Transcription Repression Activity by Multiple Mechanisms

Drosophila melanogaster Casein Kinase II Interacts with and Phosphorylates the Basic Helix-Loop-Helix Proteins m5, m7, and m8 Derived from the Enhancer of split Complex

Contact Info

Product

Resources

About

Casein kinase II phosphorylates Ser⁴⁶⁸ in the PEST domain of the Drosophila IκB homologue cactus