Serine/threonine-protein kinase A-Raf, or simplyA-Raf, is anenzyme that in humans is encoded by theARAFgene.[5] It belongs to theRaf kinase family ofserine/threonine-specific protein kinases, which also includes Raf-1 and B-Raf.[6] A-Raf is involved in theMAPK/ERK pathway, where it contributes to cell signaling processes that regulate proliferation, survival, and differentiation. Compared to Raf-1 and B-Raf, A-Raf is less well studied and exhibits distinct structural and regulatory features, including low kinase activity and alternative splicing in cancer. In addition to its role in MAPK signaling, A-Raf has functions in apoptosis suppression, cancer metabolism, and endocytic trafficking.
A-Raf, a member of the Raf kinase family, shares a conserved domain architecture with B-Raf and C-Raf, comprising three conserved regions: CR1, CR2, and CR3.
CR1 (Conserved Region 1): This N-terminal region contains the Ras-binding domain (RBD) and the cysteine-rich domain (CRD). The RBD facilitates interaction with activated Ras-GTP, anchoring A-Raf to the plasma membrane.[7] The CRD, characterized by its zinc-binding motif, contributes to membrane association and protein-protein interactions[8] Structural studies confirm the RBD and CRD function as a single entity during Ras binding.[9]
CR2 (Conserved Region 2): Positioned between CR1 and CR3, CR2 is a serine/threonine-rich regulatory segment containing phosphorylation sites (e.g., Ser259 in Raf-1) that modulate A-Raf's activity and interactions with 14-3-3 proteins.[10] This region is critical for autoinhibition and activation dynamics.[11]
CR3 (Conserved Region 3): The C-terminal kinase domain exhibits the bilobal architecture characteristic of protein kinases, with an ATP-binding site between the N-terminal and C-terminal lobes.[12] Structural analyses reveal similarities to tyrosine kinase-like (TKL) group members[13]
The RBD adopts a ubiquitin-like fold critical for Ras-GTP interaction.[14], while the CRD's zinc-binding motif stabilizes membrane association.[15] A-Raf's activity is regulated by phosphorylation-dependent 14-3-3 binding.[16] and isoform dimerization, which is essential for MAPK pathway activation.[17][18]
A-Raf shares the canonical role of Raf kinases in the MAPK signaling cascade. Upon activation by Ras, A-Raf translocates from the cytosol to the plasma membrane, where it phosphorylates and activates MEK proteins. This activation leads to downstream ERK signaling and promotes cell cycle progression and proliferation.[19]
Among the Raf isoforms, A-Raf exhibits the lowest kinase activity toward MEK proteins.[20] This may be due to amino acid substitutions in a negatively charged region upstream of the kinase domain (the N-region), which result in low basal activity.[21]
A-Raf is also the only Raf kinase known to be regulated by steroid hormones.[22] In its inactive form, A-Raf is bound to 14-3-3 proteins in the cytosol; activation by Ras causes its translocation to the plasma membrane.
Beyond the MAPK pathway, A-Raf has additional functions. It inhibits MST2, a proapoptotic kinase, thereby suppressing apoptosis. This inhibitory activity is dependent on the expression of full-length A-Raf protein, which is maintained by the splicing factor hnRNP H.[23]
A-Raf also regulates energy metabolism by interacting with pyruvate kinase M2 (PKM2), a key enzyme in cancer cell glycolysis. By promoting a conformational shift from the dimeric to the tetrameric form of PKM2, A-Raf enhances its enzymatic activity and shifts glucose utilization from biosynthesis toward energy production.[24]
In addition, A-Raf has been implicated in endocytic membrane trafficking. Upon activation by receptor tyrosine kinases and Ras, A-Raf localizes to phosphatidylinositol 4,5-bisphosphate (PtdIns(4,5)P2)-rich membranes and signals to endosomes, leading to activation of ARF6, a key regulator of endocytosis.[25]
A-Raf may contribute to tumorigenesis through multiple mechanisms. In cancer cells, overexpression of hnRNP H enhances the production of full-length A-Raf, which inhibits MST2 and prevents apoptosis. The downregulation of hnRNP H, in contrast, leads to alternative splicing of theARAF gene and loss of this anti-apoptotic activity.[26]
A-Raf's regulation of PKM2 activity further links it to cancer metabolism. By promoting glycolytic flux toward pyruvate and lactate production, A-Raf may help sustain the high energy demands of rapidly proliferating tumor cells.[27]
Because A-Raf modulates both apoptosis and metabolism—two critical hallmarks of cancer—it may represent a potential target for future cancer therapies.
^Defrise M, Kinahan PE, Townsend DW, Michel C, Sibomana M, Newport DF (April 1997). "Exact and approximate rebinning algorithms for 3-D PET data".IEEE Transactions on Medical Imaging.16 (2):145–158.doi:10.1109/42.563660.PMID9101324.
^Defrise M, Kinahan PE, Townsend DW, Michel C, Sibomana M, Newport DF (April 1997). "Exact and approximate rebinning algorithms for 3-D PET data".IEEE Transactions on Medical Imaging.16 (2):145–158.doi:10.1109/42.563660.PMID9101324.
^Motegi A, Fujimoto J, Kotani M, Sakuraba H, Yamamoto T (July 2004). "ALK receptor tyrosine kinase promotes cell growth and neurite outgrowth".Journal of Cell Science.117 (Pt 15):3319–3329.doi:10.1242/jcs.01183.PMID15226403.
^Rimmer A (June 2018). "Overseas doctors must not be used just to fill rota gaps, says leading consultant".BMJ.361 k2654.doi:10.1136/bmj.k2654.PMID29907696.
^abcdeYuryev A, Wennogle LP (February 2003). "Novel raf kinase protein-protein interactions found by an exhaustive yeast two-hybrid analysis".Genomics.81 (2):112–125.doi:10.1016/S0888-7543(02)00008-3.PMID12620389.
^Yin XL, Chen S, Gu JX (February 2002). "Identification of TH1 as an interaction partner of A-Raf kinase".Molecular and Cellular Biochemistry.231 (1–2):69–74.doi:10.1023/A:1014437024129.PMID11952167.S2CID19362635.
Papin C, Eychène A, Brunet A, Pagès G, Pouysségur J, Calothy G, et al. (1995). "B-Raf protein isoforms interact with and phosphorylate Mek-1 on serine residues 218 and 222".Oncogene.10 (8):1647–1651.PMID7731720.
Fang Y, Johnson LM, Mahon ES, Anderson DH (2002). "Two phosphorylation-independent sites on the p85 SH2 domains bind A-Raf kinase".Biochemical and Biophysical Research Communications.290 (4):1267–1274.Bibcode:2002BBRC..290.1267F.doi:10.1006/bbrc.2002.6347.PMID11812000.
Yuryev A, Wennogle LP (2003). "Novel raf kinase protein-protein interactions found by an exhaustive yeast two-hybrid analysis".Genomics.81 (2):112–125.doi:10.1016/S0888-7543(02)00008-3.PMID12620389.
