The solution structure of the cytokine-binding domain of the common β-chain of the receptors for granulocyte-macrophage colony-stimulating factor, interleukin-3 and interleukin-5 1 1Edited by P. E. Wright

2000 ◽  
Vol 297 (4) ◽  
pp. 989-1001 ◽  
Author(s):  
Terrence D Mulhern ◽  
Angel F Lopez ◽  
Richard J D’Andrea ◽  
Craig Gaunt ◽  
Lucianne Vandeleur ◽  
...  
Keyword(s):  
Solution Structure ◽  
Binding Domain ◽  
Colony Stimulating Factor ◽  
Interleukin 5 ◽  
Interleukin 3 ◽  
Macrophage Colony Stimulating Factor ◽  
The Common ◽  
Macrophage Colony ◽  
Β Chain ◽  
Stimulating Factor

Identification of a 14-3-3 Binding Sequence in the Common β Chain of the Granulocyte-Macrophage Colony-Stimulating Factor (GM-CSF), Interleukin-3 (IL-3), and IL-5 Receptors That Is Serine-Phosphorylated by GM-CSF

Blood ◽  
1999 ◽  
Vol 94 (6) ◽  
pp. 1933-1942 ◽  
Author(s):  
F.C. Stomski ◽  
M. Dottore ◽  
W. Winnall ◽  
M.A. Guthridge ◽  
J. Woodcock ◽  
...  
Keyword(s):  
Binding Site ◽  
Signaling Molecules ◽  
Colony Stimulating Factor ◽  
Interleukin 3 ◽  
Gm Csf ◽  
Macrophage Colony Stimulating Factor ◽  
The Common ◽  
Macrophage Colony ◽  
Β Chain ◽  
Stimulating Factor

Abstract The common β chain (βc) of the granulocyte-macrophage colony-stimulating factor (GM-CSF), interleukin-3 (IL-3), and IL-5 receptors is the major signaling subunit of these receptors coupling ligand binding to multiple biological activities. It is thought that these multiple functions arise as a consequence of the recruitment of specific signaling molecules to tyrosine-phosphorylated residues in the cytoplasmic domain of βc. However, the contribution of serine phosphorylation in βc to the recruitment of signaling molecules is not known. We show here the identification of a phosphoserine motif in the cytoplasmic domain of βc that interacts with the adaptor protein 14-3-3ζ. Coimmunoprecipitation and pull-down experiments with a glutathione S-transferase (GST):14-3-3ζ fusion protein showed that 14-3-3 directly associates with βc but not the GM-CSF receptor  chain. C-terminal truncation mutants of βcfurther showed that a region between amino acids 544 and 626 in βc was required for its association with 14-3-3ζ. This region contains the sequence 582HSRSLP587, which closely resembles the RSXSXP (where S is phosphorylated) consensus 14-3-3 binding site identified in a number of signaling molecules, including Raf-1. Significantly, substitution of582HSRSLP587 for EFAAAA completely abolished interaction of βc with GST–14-3-3ζ. Furthermore, the interaction of βc with GST–14-3-3 was greatly reduced in the presence of a peptide containing the 14-3-3 binding site, but only when 585Ser was phosphorylated. Direct binding experiments showed that the peptide containing phosphorylated 585Ser bound 14-3-3ζ with an affinity of 150 nmol/L. To study the regulation of 585S phosphorylation in vivo, we raised antibodies that specifically recognized 585Ser-phosphorylated βc. Using these antibodies, we showed that GM-CSF stimulation strongly upregulated 585Ser phosphorylation in M1 myeloid leukemic cells. The proximity of the SHC-binding site (577Tyr) to the 14-3-3–binding site (582HSRSLP587) and their conservation between mouse, rat, and human βc but not in other cytokine receptors suggest that they form a distinct motif that may subserve specialized functions associated with the GM-CSF, IL-3, and IL-5 receptors.

Identification of a 14-3-3 Binding Sequence in the Common β Chain of the Granulocyte-Macrophage Colony-Stimulating Factor (GM-CSF), Interleukin-3 (IL-3), and IL-5 Receptors That Is Serine-Phosphorylated by GM-CSF

Blood ◽  
1999 ◽  
Vol 94 (6) ◽  
pp. 1933-1942 ◽  
Author(s):  
F.C. Stomski ◽  
M. Dottore ◽  
W. Winnall ◽  
M.A. Guthridge ◽  
J. Woodcock ◽  
...  
Keyword(s):  
Binding Site ◽  
Signaling Molecules ◽  
Colony Stimulating Factor ◽  
Interleukin 3 ◽  
Gm Csf ◽  
Macrophage Colony Stimulating Factor ◽  
The Common ◽  
Macrophage Colony ◽  
Β Chain ◽  
Stimulating Factor

The common β chain (βc) of the granulocyte-macrophage colony-stimulating factor (GM-CSF), interleukin-3 (IL-3), and IL-5 receptors is the major signaling subunit of these receptors coupling ligand binding to multiple biological activities. It is thought that these multiple functions arise as a consequence of the recruitment of specific signaling molecules to tyrosine-phosphorylated residues in the cytoplasmic domain of βc. However, the contribution of serine phosphorylation in βc to the recruitment of signaling molecules is not known. We show here the identification of a phosphoserine motif in the cytoplasmic domain of βc that interacts with the adaptor protein 14-3-3ζ. Coimmunoprecipitation and pull-down experiments with a glutathione S-transferase (GST):14-3-3ζ fusion protein showed that 14-3-3 directly associates with βc but not the GM-CSF receptor  chain. C-terminal truncation mutants of βcfurther showed that a region between amino acids 544 and 626 in βc was required for its association with 14-3-3ζ. This region contains the sequence 582HSRSLP587, which closely resembles the RSXSXP (where S is phosphorylated) consensus 14-3-3 binding site identified in a number of signaling molecules, including Raf-1. Significantly, substitution of582HSRSLP587 for EFAAAA completely abolished interaction of βc with GST–14-3-3ζ. Furthermore, the interaction of βc with GST–14-3-3 was greatly reduced in the presence of a peptide containing the 14-3-3 binding site, but only when 585Ser was phosphorylated. Direct binding experiments showed that the peptide containing phosphorylated 585Ser bound 14-3-3ζ with an affinity of 150 nmol/L. To study the regulation of 585S phosphorylation in vivo, we raised antibodies that specifically recognized 585Ser-phosphorylated βc. Using these antibodies, we showed that GM-CSF stimulation strongly upregulated 585Ser phosphorylation in M1 myeloid leukemic cells. The proximity of the SHC-binding site (577Tyr) to the 14-3-3–binding site (582HSRSLP587) and their conservation between mouse, rat, and human βc but not in other cytokine receptors suggest that they form a distinct motif that may subserve specialized functions associated with the GM-CSF, IL-3, and IL-5 receptors.

scholarly journals A Truncated Isoform of the Human β Chain Common to the Receptors for Granulocyte-Macrophage Colony-Stimulating Factor, Interleukin-3 (IL-3), and IL-5 With Increased mRNA Expression in Some Patients With Acute Leukemia

Blood ◽  
1998 ◽  
Vol 91 (1) ◽  
pp. 54-63 ◽  
Author(s):  
Rosemary E. Gale ◽  
Robin W. Freeburn ◽  
Asim Khwaja ◽  
Rajesh Chopra ◽  
David C. Linch
Keyword(s):  
Amino Acids ◽  
Acute Leukemia ◽  
Myeloid Cells ◽  
Colony Stimulating Factor ◽  
Interleukin 3 ◽  
Gm Csf ◽  
Macrophage Colony Stimulating Factor ◽  
Macrophage Colony ◽  
Β Chain ◽  
Stimulating Factor

We report here a naturally occurring isoform of the human β chain common to the receptors for granulocyte-macrophage colony-stimulating factor (GM-CSF), interleukin-3 (IL-3), and IL-5 (GMRβC) with a truncated intracytoplasmic tail caused by deletion of a 104-bp exon in the membrane-proximal region of the chain. This β intracytoplasmic truncated chain (βIT) has a predicted tail of 46 amino acids, instead of 432 for βC, with 23 amino acids in common with βC and then a new sequence of 23 amino acids. In primary myeloid cells, βIT comprised approximately 20% of the total β chain message, but was increased up to 90% of total in blast cells from a significant proportion of patients with acute leukemia. Specific anti-βITantibodies demonstrated its presence in primary myeloid cells and cell lines. Coexpression of βIT converted low-affinity GMRα chains (KD 2.5 nmol/L) to higher-affinity αβ complexes (KD 200 pmol/L). These could bind JAK2 that was tyrosine-phosphorylated by stimulation with GM-CSF. βITdid not support GM-CSF–induced proliferation when cotransfected with GMRα into CTLL-2 cells. Therefore, it may interfere with the signal-transducing properties of the βC chain and play a role in the pathogenesis of leukemia.

scholarly journals A mutation of the common receptor subunit for interleukin-3 (IL-3), granulocyte-macrophage colony-stimulating factor, and IL-5 that leads to ligand independence and tumorigenicity

Blood ◽  
1994 ◽  
Vol 83 (10) ◽  
pp. 2802-2808 ◽  
Author(s):  
R D'Andrea ◽  
J Rayner ◽  
P Moretti ◽  
A Lopez ◽  
GJ Goodall ◽  
...  
Keyword(s):  
Large Family ◽  
Mutant Form ◽  
Sequence Motifs ◽  
Colony Stimulating Factor ◽  
Interleukin 5 ◽  
Interleukin 3 ◽  
Conserved Sequence ◽  
Macrophage Colony Stimulating Factor ◽  
Macrophage Colony ◽  
Stimulating Factor

Abstract The cytokines interleukin-3, interleukin-5, and granulocyte-macrophage colony-stimulating factor bind with high affinity to a receptor complex that contains a ligand-specific alpha-chain and a common beta-chain, h beta c. We report here the isolation of a mutant form of h beta c, from growth factor-independent cells, that arose spontaneously after infection of a murine factor-dependent hematopoietic cell line (FDC-P1) with a retroviral h beta c expression construct. Analysis of this h beta c mutation shows that a small (37 amino acid) duplication of extracellular sequence that includes two conserved sequence motifs is sufficient to confer ligand-independent growth on these cells and lead to tumourigenicity. Because this is a conserved region in the cytokine receptor superfamily, our results suggest that the large family of cytokine receptors has the capacity to become oncogenically active.

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