Nucleotide binding domain and leucine-rich repeat pyrin domain-containing protein 12: characterization of its binding to hematopoietic cell kinase.

Abstract

Protein-protein interactions are key to define the function of nucleotide binding domain (NBD) and leucine-rich repeat (LRR) family, pyrin domain (PYD)-containing protein 12 (NLRP12). cDNA encoding the human PYD + NBD of NLRP12 was used as bait in a yeast two-hybrid screen with a human leukocyte cDNA library as prey. Hematopoiesis cell kinase (HCK), a member of the c-SRC family of non-receptor tyrosine kinases, was among the top hits. The C-terminal 40 amino acids of HCK selectively bound to NLRP12's PYD + NBD, but not to that of NLRP3 and NLRP8. Amino acids F503, I506, Q507, L510, and D511 of HCK are critical for the binding of HCK's C-terminal 40 amino acids to NLRP12's PYD + NBD. Additionally, the C-terminal 30 amino acids of HCK are sufficient to bind to NLRP12's PYD + NBD, but not to its PYD alone nor to its NBD alone. In cell lines that express HCK endogenously, it was co- immunoprecipitated with stably expressed exogenous NLRP12. Also, NLRP12 co-immunoprecipitated and co-localized with HCK when both were overexpressed in 293T cells. In addition, in this overexpression system, steady-state NLRP12 protein expression levels significantly decreased when HCK was co-expressed. Bioinformatic analysis showed that HCK mRNA co-occurred with NLRP12 mRNA, but not with other NLRP mRNAs, in blood and marrow samples from acute myeloid leukemia (AML) patients. The mRNA of NLRP12 is also co-expressed with HCK in AML patient samples, and the levels of mRNA expression of each are correlated. Together these data suggest that NLRP12, through its binding to HCK, may have an effect on the pathogenesis of AML.

• Authors: Yue Zhang, Curtis T Okamoto
• Journal: International journal of biological sciences (Int J Biol Sci) Vol. 16 Issue 9 Pg. 1507-1525 ( 2020) ISSN: 1449-2288 [Electronic] Australia
• PMID: 32226298 (Publication Type: Journal Article)
• Copyright: © The author(s).

Chemical References

• Intracellular Signaling Peptides and Proteins
• NLRP12 protein, human
• Proto-Oncogene Proteins c-hck

Topics

• Bone Marrow (metabolism)
• Cell Line
• HEK293 Cells
• Humans
• Intracellular Signaling Peptides and Proteins (blood, chemistry, genetics, metabolism)
• Leukemia, Myeloid, Acute (blood, genetics, metabolism)
• Protein Binding
• Protein Interaction Domains and Motifs
• Proto-Oncogene Proteins c-hck (blood, chemistry, genetics, metabolism)
• Two-Hybrid System Techniques