Molecular Basis of Oncogenic PI3K Proteins

Abstract

The dysregulation of phosphatidylinositol 3-kinase (PI3K) signaling plays a pivotal role in driving neoplastic transformation by promoting uncontrolled cell survival and proliferation. This oncogenic activity is primarily caused by mutations that are frequently found in PI3K genes and constitutively activate the PI3K signaling pathway. However, tumorigenesis can also arise from nonmutated PI3K proteins adopting unique active conformations, further complicating the understanding of PI3K-driven cancers. Recent structural studies have illuminated the functional divergence among highly homologous PI3K proteins, revealing how subtle structural alterations significantly impact their activity and contribute to tumorigenesis. In this review, we summarize current knowledge of Class I PI3K proteins and aim to unravel the complex mechanism underlying their oncogenic traits. These insights will not only enhance our understanding of PI3K-mediated oncogenesis but also pave the way for the design of novel PI3K-based therapies to combat cancers driven by this signaling pathway.

Keywords: PI3K; oncogenic mutation; oncogenic transformation; protein structure.

Figure 1

Lipid phosphorylation by PI3K kinases and its consequences. At the catalytic center of the Class I PI3K kinases—p110α (blue), p110β (red), p110δ (green), or p110γ (purple)—the γ-phosphate from ATP is transferred to the D3 position of the inositol ring of PIP2 (arrow), yielding PIP3 and ADP. PIP3 subsequently recruits and activates the PH effector PDK1, leading to the activation of AKT. Once activated, AKT changes the following cellular processes: (1) Stimulating cell proliferation via activating MTOR; (2) Promoting cell growth by inactivating glycogen synthase kinase 3β (GSK3β) to release its inhibition of Cyclin D; (3) Suppressing cell death via enhancing the activity of anti-apoptotic factors (e.g., MCL1/BCL2) while inhibiting pro-apoptotic proteins (e.g., BH3-only proteins). The activation of cell survival, proliferation, and growth by PI3K activation ultimately contributes to oncogenic transformation. The structural models of human PI3K kinases were predicted using AlphaFold. The structures of PIP2, PIP3, ADP, and ATP were retrieved from PubChem. All 3D structures were reconstructed using ChimeraX-1.8, and the catalytic centers of the four Class I kinases were aligned using ChimeraX-1.8 Matchmaker.

Figure 2

Alignment of Class I PI3K kinases. Protein sequences of PI3K kinases were obtained from UniProt: PIK3CA/p110α: P42336; PIK3CB/p110β: P42338; PIK3CD/p110δ: O00329; PIK3CG/p110γ: P48736. Sequence alignment was performed using Clustal Omega and COBALT. Structural and functional domains were annotated based on UniProt and published literature (PMID:10580505; 18633356; 21362552; 34452907). Functional domains are color-coded: ABD (dark blue), RBD (light blue), C2 (cyan), HD (green), KD (red), and Linker regions (grey). Domains and motifs in p110α (representing class IA) are shown on the top, while those in p110γ (IB) are shown on the bottom. Identical residues are highlighted in red (all p110 kinases), blue (class IA p110 kinases), or green (shared between p110β and p110δ). Conserved residues are shown in magenta. Structural elements such as β-sheets (arrows), α-helices (cylinders), and loops/turns (lines) are indicated with undetermined regions represented by dotted lines. Key motifs, including nuclear localization signal, ATP-binding sites, catalytic loop, and activation loop, are labeled. Oncogenic mutations are marked with an asterisk.

Figure 2

Alignment of Class I PI3K kinases. Protein sequences of PI3K kinases were obtained from UniProt: PIK3CA/p110α: P42336; PIK3CB/p110β: P42338; PIK3CD/p110δ: O00329; PIK3CG/p110γ: P48736. Sequence alignment was performed using Clustal Omega and COBALT. Structural and functional domains were annotated based on UniProt and published literature (PMID:10580505; 18633356; 21362552; 34452907). Functional domains are color-coded: ABD (dark blue), RBD (light blue), C2 (cyan), HD (green), KD (red), and Linker regions (grey). Domains and motifs in p110α (representing class IA) are shown on the top, while those in p110γ (IB) are shown on the bottom. Identical residues are highlighted in red (all p110 kinases), blue (class IA p110 kinases), or green (shared between p110β and p110δ). Conserved residues are shown in magenta. Structural elements such as β-sheets (arrows), α-helices (cylinders), and loops/turns (lines) are indicated with undetermined regions represented by dotted lines. Key motifs, including nuclear localization signal, ATP-binding sites, catalytic loop, and activation loop, are labeled. Oncogenic mutations are marked with an asterisk.

Figure 3

Alignment of Class I PI3K regulatory subunits. Protein sequences of PI3K regulatory subunits were obtained from UniProt: PIK3R1/p85α/p55α/p50α: P27986; PIK3R2/p85β: O00459; PIK3R3/p55γ: O92569; PIK3R5/p101: Q8WYR; PIK3R6/p84: Q5UE93. Sequence alignment was conducted using Clustal Omega and COBALT. Structural and functional domains were annotated based on Uniprot, AlphaFold-predicted 3D structures, and published literature (PMID:21362552 and 35429500). Functional domains in IA adapters are color-coded: SH3 (brown), P1/P2 (black), BH (brown), nSH2 (brown), iSH2 (dark brown), and cSH2 (brown). Functional domains in IB adapters are also color-coded: PBD/GBD (brown). Other N-terminal or C-terminal secondary structures are labeled in grey. Identical residues are highlighted in red among all Class IA adapters (p85α, p85β, p55γ, p55α and p50α) or between the Class IB adapters (p101 and p84). Identical residues are highlighted in green between p85α and p85β. Conserved residues among all adapters are in magenta. Structural elements are represented as follows: β-sheets (arrows), α-helices (cylinders), and loops/turns (lines).

Figure 3

Alignment of Class I PI3K regulatory subunits. Protein sequences of PI3K regulatory subunits were obtained from UniProt: PIK3R1/p85α/p55α/p50α: P27986; PIK3R2/p85β: O00459; PIK3R3/p55γ: O92569; PIK3R5/p101: Q8WYR; PIK3R6/p84: Q5UE93. Sequence alignment was conducted using Clustal Omega and COBALT. Structural and functional domains were annotated based on Uniprot, AlphaFold-predicted 3D structures, and published literature (PMID:21362552 and 35429500). Functional domains in IA adapters are color-coded: SH3 (brown), P1/P2 (black), BH (brown), nSH2 (brown), iSH2 (dark brown), and cSH2 (brown). Functional domains in IB adapters are also color-coded: PBD/GBD (brown). Other N-terminal or C-terminal secondary structures are labeled in grey. Identical residues are highlighted in red among all Class IA adapters (p85α, p85β, p55γ, p55α and p50α) or between the Class IB adapters (p101 and p84). Identical residues are highlighted in green between p85α and p85β. Conserved residues among all adapters are in magenta. Structural elements are represented as follows: β-sheets (arrows), α-helices (cylinders), and loops/turns (lines).

Figure 4

Functional domains in Class I PI3K proteins. (A) Illustration of functional domains in Class I PI3K kinases. (B) Illustration of functional domains in Class I PI3K adapters. Locations of domains are indicated. Linker regions are shown as grey bars. *: mutated residue.

Figure 5

Homology among PI3K proteins. Using the sequence alignments shown in Figure 2 and Figure 3, homologous amino acids were identified, and the percentage of homology was calculated by dividing the number of homologous residues by the total number of residues in each PI3K kinases or adapters. Shown are homologies of full-length p110 kinase and their functional domains (A) and homologies of adapters and their functional domains (B).

Figure 6

PI3K 3D structures. Since published crystal structures are often derived from truncated proteins or use non-human proteins, we employed AlphaFold to predict 3D conformations of full-length human PI3K proteins. These structures were aligned using ChimeraX-1.8 Matchmaker. (A) Aligned 3D structures of full-length PI3K kinases, which are color-coded as follows: p110α (blue), p110β (red), p110δ (green), p110γ (purple), and linker regions (light grey). (B) Aligned 3D structures of Class IA adaptors, which are color-coded as follows: p85α (grey), p85β (lime), and p55γ (gold). (C) Aligned 3D structures of Class IB adaptors, which are color-coded as follows: p101 (magenta) and p84 (light blue). Functional domains, as illustrated in Figure 2, Figure 3 and Figure 4, are labeled. The aligned structures are shown with a 180° rotation for a comprehensive view.

Figure 7

Structural homology of PI3K proteins and domains. RMSDs of full-length PI3K kinases or adapters, along with their functional domains, were determined using ChimeraX-1.8 Matchmaker. These values were plotted against percentages of homology shown in Figure 3. PI3K proteins are color-coded as follows: p110α (blue); p110β (red), p110δ (green); p110γ (purple); linker regions (light grey); p85α (grey); p85β (lime); p55γ (gold); p101 (magenta); and p84 (light blue). (A) Kinases and their domains/motifs. (B) Adaptors and their domains/motifs. Regions with high similarities (sequence homology > 50%/RMSD < 4Å) are highlighted in green, whereas low similarities (sequence homology < 50%/RMSDs > 4Å) are indicated in yellow. Red blank circles: Class IA kinases p110α/β/δ vs. IB kinase p110γ; Blue blank rectangles: within Class IA kinases p110α/β/δ; Purple filled triangles: p110α vs. p110β (α/β); Green blank diamonds: p110α vs. p110δ (α/δ); Cyan filled hexagons: p110β vs. p110δ (β/δ); Brown Half-filled rectangles: within full-length Class IA adaptors p85α/p85β/p55γ; Magenta inverted blank triangles: p85α vs. p85β; Black filled diamonds: full length p101 vs. full length p84; Asterisks: domains in p101 vs. domains in p84.

Figure 8

Intra- and inter-subunit interactions in PI3K protein complexes. 3D structures of PI3K protein complexes were retrieved from the PDB database and analyzed using ChimeraX-1.8. Full structures are displayed in the left panel, while the right panel highlights enlarged structures with detailed interface interactions. The interactions include: (A) Between p110α’s ABD and p85α’s iSH2; (B) Between p110γ’s ABD and RBD C2 linker; (C) Between p110γ’s RBD and RAS; (D) Between p110α’s C2 and p85α’s iSH2; (E) Between p110γ’s C2 and p101′s PBD; (F) Between p110α’s HD and p110α’s C2/KD/p85α’s nSH2; (G) Between p110γ’s HD and p110γ’s C2/KD/p101. PDB access numbers for representative structures are provided.

Figure 9

Mutations in PI3K genes in cancer. Data were retrieved from cBioportal, encompassing 800,085 tumor samples across 149 tumor types from 224 studies. (A) Mutation frequencies of PI3K genes. The figure shows the number of samples with or without mutations in PI3K genes as well as mutation frequencies. Mutation frequencies were calculated by dividing the number of samples with PI3K mutations with the total number of samples. (B) Driver mutations in PI3K genes. Shown are numbers of mutations either recognized as driver mutations or variants of uncertain significance. Frequencies of driver mutations, which were obtained by dividing the number of driver mutations by the total number of mutations, are also shown.

Figure 10

Hotspot mutations in PI3K proteins. Data were retrieved from cBioportal, Mutation plots for PI3K kinases (A) or adaptors (B) were extracted from cBioportal and replotted. Shown are hotspot mutations in functional domains of p110α, p110β, p110δ, p110γ, p85α, or p85β. *: stop-codon. Dark colors: driver mutations; Light colors: variants of uncertain significance; Dark or light green dots: missense mutations; Black or grey dots: truncations; Dark or light brown dots: in-frame mutations; Dark or light orange dots: splice mutations; Dark or light purple dots: fusion mutations.