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Cited by 11Biochemical and Molecular ResearchRNA and protein synthesis mechanismsGenomics and Phylogenetic Studies

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The role of gene duplication and paralog specialisation in the evolution of the mammalian PRPS complex

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Karki, Bibek R., MacMillan, Austin C., Vicente-Muñoz, Sara, Greis, Kenneth D., Romick, Lindsey E., Cunningham, John T.·2025·Nature Communications
DOI 10.1038/s41467-025-61216-zPMC PMC12238573

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Abstract

The phosphoribosyl pyrophosphate synthetase (PRPS) enzyme catalyzes a chokepoint reaction in nucleotide production, making it essential for life. Here, we show that the presence of multiple PRPS-encoding genes is a hallmark trait of eukaryotes, and we find that gains or losses of paralogs are associated with major branching events in the eukaryotic tree. We pinpoint the evolutionary origins and define the individual roles for each of the mammalian PRPS paralogs, which we demonstrate work together as a heterogeneous multicomponent complex. Employing isogenic cells representing all viable individual or combinatorial assembly states, we dissect the basic organizational principles of the enzyme complex and characterize the emergent properties responsible for paralog specialization, including new modes of regulation that govern complex assembly and activity in vivo. Collectively, our study demonstrates how evolution has transformed a single PRPS enzyme into a biochemical complex endowed with novel functional and regulatory features that fine-tune mammalian metabolism.

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Fig. 1 Loss of Class II PRPS associated with expanded Class I PRPS homolog repertoire. A Phylogenetic distribution of PRPS homologs in eukaryotes, with presence/absence of NHR-containing PRPS homologs in opisthokonts. PRPSAP2 denotes orthologs of mammalian PRPSAP2; Prs1 and Prs5 represent S. cerevisiae Prs1 and Prs5, respectively. Black asterisk indicates Class II presence across most metazoans, excluding Craniata. Red asterisks indicate Class II PRPS identified in only two Nuclearia species and one Chytridomycota species. B Conserved splice site junctions among PRPS homologs across different representative organisms in Opisthokonta. Gene structures for PRPS, PRPSAP2, and Prs5-encoding genes shown with exons as colored boxes; introns not displayed. Multiple sequence alignment (MSA) of translated sequences via Clustal Omega highlights a conserved splice site junction (red arrow) with adjacent amino acids shown. Top bar shows full H. sapiens PRPS1 with RF loop (red box) and CF loop (black box). Dotted red and black lines project corresponding RF and CF loops positions, respectively onto other homologs. Insertions in RF and CF loops of PRPSAP2 and Prs5 shown with red and black hatch marks, respectively (hatch marks not to scale; NHRs vary in length). C Phylogenetic distribution of PRPS homologs in chordates. Mostly present – indicates present in most taxa. Black asterisk highlights that Class II PRPS is found in most urochordates, except Oikopleura, which has additional Class I PRPS homologs not observed in other organisms from this clade. D, E Conserved splice site junctions between PRPS1 and PRPS2 (D), and PRPSAP2 and PRPSAP1 (E) across different representative organisms in jawed Vertebrata. Gene structures for PRPS, PRPSAP2, and Prs5-encoding genes shown with exons as colored boxes; introns not displayed. Dotted red and black lines denote RF and CF loop regions, respectively; insertions in PRPSAP2 and PRPSAP1 shown as red and black hatch marks, respectively (hatch marks not to scale; NHRs vary in length).
Fig. 1 Loss of Class II PRPS associated with expanded Class I PRPS homolog repertoire. A Phylogenetic distribution of PRPS homologs in eukaryotes, with presence/absence of NHR-containing PRPS homologs in opisthokonts. PRPSAP2 denotes orthologs of mammalian PRPSAP2; Prs1 and Prs5 represent S. cerevisiae Prs1 and Prs5, respectively. Black asterisk indicates Class II presence across most metazoans, excluding Craniata. Red asterisks indicate Class II PRPS identified in only two Nuclearia species and one Chytridomycota species. B Conserved splice site junctions among PRPS homologs across different representative organisms in Opisthokonta. Gene structures for PRPS, PRPSAP2, and Prs5-encoding genes shown with exons as colored boxes; introns not displayed. Multiple sequence alignment (MSA) of translated sequences via Clustal Omega highlights a conserved splice site junction (red arrow) with adjacent amino acids shown. Top bar shows full H. sapiens PRPS1 with RF loop (red box) and CF loop (black box). Dotted red and black lines project corresponding RF and CF loops positions, respectively onto other homologs. Insertions in RF and CF loops of PRPSAP2 and Prs5 shown with red and black hatch marks, respectively (hatch marks not to scale; NHRs vary in length). C Phylogenetic distribution of PRPS homologs in chordates. Mostly present – indicates present in most taxa. Black asterisk highlights that Class II PRPS is found in most urochordates, except Oikopleura, which has additional Class I PRPS homologs not observed in other organisms from this clade. D, E Conserved splice site junctions between PRPS1 and PRPS2 (D), and PRPSAP2 and PRPSAP1 (E) across different representative organisms in jawed Vertebrata. Gene structures for PRPS, PRPSAP2, and Prs5-encoding genes shown with exons as colored boxes; introns not displayed. Dotted red and black lines denote RF and CF loop regions, respectively; insertions in PRPSAP2 and PRPSAP1 shown as red and black hatch marks, respectively (hatch marks not to scale; NHRs vary in length).CC BY · PubMed Central
Fig. 2 Convergent evolution of PRPSAP-like homologs in Amorphea. A Structure of dimeric human PRPS1 (PDB: 2HCR), with a zoom-in of catalytic site highlighting metal binding site (Cd2+), AMP (represents AMP moiety of ATP), SO42− (represents 5′-phosphate of R5P), and several conserved active site residues (magenta). D171 coordinates metal binding, K194 interacts with ATP, R196, and T225 interact with R5P, and N200 stabilizes the catalytic loop. B WebLogo depicting MSA of active and regulatory site residues from PRPS1 and PRPSAP2 from representative organisms in opisthokonts (n = 44 each). Numbers below indicate corresponding residue positions in human PRPS1 (NP_002755.1) and PRPSAP2 (NP_001340030.1). Asterisks denote residues conserved in PRPS but substituted in PRPSAP2 (also shown in A). C AlphaFold2-predicted structure of human PRPSAP2 (NP_001340030.1), with a zoom-in highlighting four non-conserved residues (magenta) at corresponding active site positions in PRPS1 shown in (A). AMP modeled to indicate putative ATP binding site. D Trimeric structure of human PRPS1 (PDB: 2HCR). Red and magenta residues in dashed box represent dimer interface residues in bent (B, C) and parallel (A, B) dimers, respectively. Amino acid sequence of B. subtilis PRPS aligned with Opisthokonta PRPS homologs to identify corresponding dimer interface residues for WebLogo. Representative sequences include PRPS1 (n = 44), PRPSAP2 (n = 44) from opisthokonts, and PRPS2 (n = 46), PRPSAP1 (n = 92) from jawed vertebrates. Residue numbers based on human PRPS1 (NP_002755.1), PRPS2 (NP_002756.1), PRPSAP1 (AAH09012.1), and PRPSAP2 (NP_001340030.1). E PRPS paralogs from S. cerevisiae, H. sapiens, R. potamoides, T. trahens, and D. rotans showing relative NHR positions in paralogs with expanded CF and/or RF loops. Open bar for each representative species represents full polypeptide sequences of their ancestral PRPS (bold). Amino acid positions corresponding to RF and CF loops labeled within bars. NHR insertions sites for Prs1, Prs5, PRPSAP1, PRPSAP2, PrsB, and PRPSAP-like homologs marked by triangles above/below bars. Numbers next to triangles indicate amino acid count per NHR.
Fig. 2 Convergent evolution of PRPSAP-like homologs in Amorphea. A Structure of dimeric human PRPS1 (PDB: 2HCR), with a zoom-in of catalytic site highlighting metal binding site (Cd2+), AMP (represents AMP moiety of ATP), SO42− (represents 5′-phosphate of R5P), and several conserved active site residues (magenta). D171 coordinates metal binding, K194 interacts with ATP, R196, and T225 interact with R5P, and N200 stabilizes the catalytic loop. B WebLogo depicting MSA of active and regulatory site residues from PRPS1 and PRPSAP2 from representative organisms in opisthokonts (n = 44 each). Numbers below indicate corresponding residue positions in human PRPS1 (NP_002755.1) and PRPSAP2 (NP_001340030.1). Asterisks denote residues conserved in PRPS but substituted in PRPSAP2 (also shown in A). C AlphaFold2-predicted structure of human PRPSAP2 (NP_001340030.1), with a zoom-in highlighting four non-conserved residues (magenta) at corresponding active site positions in PRPS1 shown in (A). AMP modeled to indicate putative ATP binding site. D Trimeric structure of human PRPS1 (PDB: 2HCR). Red and magenta residues in dashed box represent dimer interface residues in bent (B, C) and parallel (A, B) dimers, respectively. Amino acid sequence of B. subtilis PRPS aligned with Opisthokonta PRPS homologs to identify corresponding dimer interface residues for WebLogo. Representative sequences include PRPS1 (n = 44), PRPSAP2 (n = 44) from opisthokonts, and PRPS2 (n = 46), PRPSAP1 (n = 92) from jawed vertebrates. Residue numbers based on human PRPS1 (NP_002755.1), PRPS2 (NP_002756.1), PRPSAP1 (AAH09012.1), and PRPSAP2 (NP_001340030.1). E PRPS paralogs from S. cerevisiae, H. sapiens, R. potamoides, T. trahens, and D. rotans showing relative NHR positions in paralogs with expanded CF and/or RF loops. Open bar for each representative species represents full polypeptide sequences of their ancestral PRPS (bold). Amino acid positions corresponding to RF and CF loops labeled within bars. NHR insertions sites for Prs1, Prs5, PRPSAP1, PRPSAP2, PrsB, and PRPSAP-like homologs marked by triangles above/below bars. Numbers next to triangles indicate amino acid count per NHR.CC BY · PubMed Central
Fig. 3 PRPS enzymes operate as a multimeric complex that attains heterogeneous configurations. A SDS-PAGE followed by Coomassie stain of eluates from GFP immunoprecipitation (IP) performed in NIH3T3 and HEK293T cells stably expressing PRPS1-GFP, PRPS2-GFP, and NES-GFP. NES (Nuclear export signal)-GFP used as control. Asterisk indicates a non-specific band in eluates. B Scatter plot from mass spectrometry (MS) runs of eluates from GFP IP in NIH3T3 (x-axis) and HEK293T cells (y-axis) stably expressing PRPS1-GFP. Axes represent square root-transformed SEQUEST HT scores of PRPS1-GFP normalized to control. PRPS1L1 isoform detected only in HEK29T cells. C Western blot analysis of size exclusion chromatography (SEC) fractions collected from NIH3T3 native whole cell lysates. Cell lysates were fractionated using a Superose 6 Increase 3.2/300 column. Immunoblots probing indicated PRPS complex members and internal standards shown. D–F Western blot analysis of SEC fractions from native tissue lysates of liver (D), kidney (E), and lung (F). Twelve-week-old male C57BL/6 mice were used. Lysates were fractionated on a Superose 6 Increase 3.2/300 column. Immunoblots probing PRPS complex members and internal standards are shown. Circular pictograms below SEC immunoblots schematize PRPS complex configurations; double circle denotes multiple copies of the protein interacting within the heteromeric complex. Coomassie staining (A) and western blot data (C–F) are representative of at least 3 biological repeats. Source data are provided as a Source Data file.
Fig. 3 PRPS enzymes operate as a multimeric complex that attains heterogeneous configurations. A SDS-PAGE followed by Coomassie stain of eluates from GFP immunoprecipitation (IP) performed in NIH3T3 and HEK293T cells stably expressing PRPS1-GFP, PRPS2-GFP, and NES-GFP. NES (Nuclear export signal)-GFP used as control. Asterisk indicates a non-specific band in eluates. B Scatter plot from mass spectrometry (MS) runs of eluates from GFP IP in NIH3T3 (x-axis) and HEK293T cells (y-axis) stably expressing PRPS1-GFP. Axes represent square root-transformed SEQUEST HT scores of PRPS1-GFP normalized to control. PRPS1L1 isoform detected only in HEK29T cells. C Western blot analysis of size exclusion chromatography (SEC) fractions collected from NIH3T3 native whole cell lysates. Cell lysates were fractionated using a Superose 6 Increase 3.2/300 column. Immunoblots probing indicated PRPS complex members and internal standards shown. D–F Western blot analysis of SEC fractions from native tissue lysates of liver (D), kidney (E), and lung (F). Twelve-week-old male C57BL/6 mice were used. Lysates were fractionated on a Superose 6 Increase 3.2/300 column. Immunoblots probing PRPS complex members and internal standards are shown. Circular pictograms below SEC immunoblots schematize PRPS complex configurations; double circle denotes multiple copies of the protein interacting within the heteromeric complex. Coomassie staining (A) and western blot data (C–F) are representative of at least 3 biological repeats. Source data are provided as a Source Data file.CC BY · PubMed Central
Fig. 4 Cells exclusively expressing PRPS1 display metabolic defects and decreased cellular proliferation. A Western blot validating CRISPR-Cas9-generated isogenic knockout cell lines. B Proliferation of NIH3T3 parental and knockout cell lines generated in (A) (n = 3 technical replicates). C Western blot validating HRASG12V-overexpression in NIH3T3 parental and P2/AP1/AP2 KO cell lines. Phospho-MAPK (Erk1/2) (T202/Y204) used as a marker for activation of signaling pathways upon HRASG12V overexpression. D Representative images from soft agar colony formation assay performed in NIH3T3 parental and P2/AP1/AP2 KO cells expressing HRASG12V. Scale bar, 500 μm. E Quantification of colonies from (D) (n = 3 experimental replicates). F Oxygen consumption rate (OCR) measured by Seahorse ATP Rate Assay in NIH3T3 parental and P2/AP1/AP2 KO cell lines (n = 8 technical replicates). G OCR measured by Seahorse mitochondrial stress tests in NIH3T3 parental and NDI1-expressing P2/AP1/AP2 KO cell lines (n = 7 technical replicates). H Proliferation of nucleoside supplemented and NDI1 expressing NIH3T3 P2/AP1/AP2 cell lines (n = 3 technical replicates). I13C6-glucose metabolic labeling performed in NIH3T3 parental and P2/AP1/AP2 KO cell lines for 30 min and 5 h. Unlabeled, 13C-labeled (30 min), and 13C-labeled (5 h) samples are represented in brown, green, and blue colors, respectively. 13C-enrichments quantified from 1H-NMR spectra (n = 3 experimental replicates). J Western blot analysis of SEC fractions collected from NIH3T3 P2/AP1/AP2 KO native whole-cell lysates. Cell lysates were fractionated on a Superose 6 Increase 3.2/300 column. In the pictogram below SEC immunoblots, the double circle with dotted inner circle denotes multiple copies of PRPS1 forming homo-oligomers. Data are represented as mean ± SD for (B, E–H) and mean ± SEM for (I). Statistical comparisons made using one way ANOVA followed by Tukey’s HSD post hoc test (E, I). Source data are provided as a Source Data file.
Fig. 4 Cells exclusively expressing PRPS1 display metabolic defects and decreased cellular proliferation. A Western blot validating CRISPR-Cas9-generated isogenic knockout cell lines. B Proliferation of NIH3T3 parental and knockout cell lines generated in (A) (n = 3 technical replicates). C Western blot validating HRASG12V-overexpression in NIH3T3 parental and P2/AP1/AP2 KO cell lines. Phospho-MAPK (Erk1/2) (T202/Y204) used as a marker for activation of signaling pathways upon HRASG12V overexpression. D Representative images from soft agar colony formation assay performed in NIH3T3 parental and P2/AP1/AP2 KO cells expressing HRASG12V. Scale bar, 500 μm. E Quantification of colonies from (D) (n = 3 experimental replicates). F Oxygen consumption rate (OCR) measured by Seahorse ATP Rate Assay in NIH3T3 parental and P2/AP1/AP2 KO cell lines (n = 8 technical replicates). G OCR measured by Seahorse mitochondrial stress tests in NIH3T3 parental and NDI1-expressing P2/AP1/AP2 KO cell lines (n = 7 technical replicates). H Proliferation of nucleoside supplemented and NDI1 expressing NIH3T3 P2/AP1/AP2 cell lines (n = 3 technical replicates). I13C6-glucose metabolic labeling performed in NIH3T3 parental and P2/AP1/AP2 KO cell lines for 30 min and 5 h. Unlabeled, 13C-labeled (30 min), and 13C-labeled (5 h) samples are represented in brown, green, and blue colors, respectively. 13C-enrichments quantified from 1H-NMR spectra (n = 3 experimental replicates). J Western blot analysis of SEC fractions collected from NIH3T3 P2/AP1/AP2 KO native whole-cell lysates. Cell lysates were fractionated on a Superose 6 Increase 3.2/300 column. In the pictogram below SEC immunoblots, the double circle with dotted inner circle denotes multiple copies of PRPS1 forming homo-oligomers. Data are represented as mean ± SD for (B, E–H) and mean ± SEM for (I). Statistical comparisons made using one way ANOVA followed by Tukey’s HSD post hoc test (E, I). Source data are provided as a Source Data file.CC BY · PubMed Central
Fig. 5 PRPSAPs are molecular scaffolds with preferential binding partners among PRPS isozymes. A, B Western blot analysis of SEC fractions collected from native whole cell lysates of NIH3T3 P2/AP2 KO cells (A) and P2/AP1/AP2 KO cells stably expressing AP1 (B). C ALFA pulldown from whole cell extracts of NIH3T3 parental and P2/AP2 KO cells transiently transfected with PRPS1-ALFA. D–G Western blot analysis of SEC fractions collected from native whole-cell lysates of NIH3T3 AP1 KO cells (D), AP1/AP2 KO cells (E), AP1 KO cells (F), and AP1/AP2 KO cells (G). H Summary comparing SEC profiles from NIH3T3 parental and CRISPR KO lines. The x-axis represents relative molecular weight of the complex, and the y-axis represents the number of PRPS complex members interacting in the complex. Cell lysates for (A, B, D, and E) were fractionated on a Superose 6 Increase 3.2/300 column, while cell lysates for (F) and (G) were fractionated on a Yarra SEC-2000 column. Circular pictograms below SEC immunoblots schematize PRPS complex configurations. Double circle denotes multiple copies of the protein interacting in a heteromeric complex. Double circle with dotted inner circle denotes multiple copies forming homo-oligomers. Single circle denotes a single protein interacting within the complex. Circle with inner vertical lines denotes proteins forming a trimer or tetramer. Western blot data (A–G) are representative of at least 2 biological repeats. Source data are provided as a Source Data file.
Fig. 5 PRPSAPs are molecular scaffolds with preferential binding partners among PRPS isozymes. A, B Western blot analysis of SEC fractions collected from native whole cell lysates of NIH3T3 P2/AP2 KO cells (A) and P2/AP1/AP2 KO cells stably expressing AP1 (B). C ALFA pulldown from whole cell extracts of NIH3T3 parental and P2/AP2 KO cells transiently transfected with PRPS1-ALFA. D–G Western blot analysis of SEC fractions collected from native whole-cell lysates of NIH3T3 AP1 KO cells (D), AP1/AP2 KO cells (E), AP1 KO cells (F), and AP1/AP2 KO cells (G). H Summary comparing SEC profiles from NIH3T3 parental and CRISPR KO lines. The x-axis represents relative molecular weight of the complex, and the y-axis represents the number of PRPS complex members interacting in the complex. Cell lysates for (A, B, D, and E) were fractionated on a Superose 6 Increase 3.2/300 column, while cell lysates for (F) and (G) were fractionated on a Yarra SEC-2000 column. Circular pictograms below SEC immunoblots schematize PRPS complex configurations. Double circle denotes multiple copies of the protein interacting in a heteromeric complex. Double circle with dotted inner circle denotes multiple copies forming homo-oligomers. Single circle denotes a single protein interacting within the complex. Circle with inner vertical lines denotes proteins forming a trimer or tetramer. Western blot data (A–G) are representative of at least 2 biological repeats. Source data are provided as a Source Data file.CC BY · PubMed Central
Fig. 6 Translation control of PRPS complex assembly. A–C Western blot analysis of SEC fractions collected from native whole-cell lysates of NIH3T3 AP1 KO cells stably expressing AP1 lacking the non-homologous region (NHR) (A), AP1/AP2 KO cells stably expressing chimeric AP1 with AP2’s N-terminus (residues 1–95) B and chimeric AP2 with AP1’s N-terminus (residues 1–95) (C). D Schematic representation of alternative start sites in mammalian AP1 and AP2 and their consequent translation into short and long isoforms. Base positions correspond to the mouse homologs. TSS1 and TSS2 represent upstream and downstream translation start sites, respectively. E Multiple isoforms for AP1 and AP2 detected under optimal SDS-PAGE resolution. Arrows in immunoblots indicate the longer isoforms of AP1 and AP2, corresponding to N-terminal leader sequences of 29 and 12 amino acids, respectively. F Phylogenetic distribution of PRPS homologs (PRPS1, PRPS2, PRPSAP2, PRPSAP2 with N-terminal leader sequence, PRPSAP1, and PRPSAP1 with N-terminal leader sequence) in chordates. Presence/absence noted across the tree. PRPSAP2 and PRPSAP1 isoforms with N-terminal leader sequences emerged in ancestors of Amniota and Osteichthyes, respectively. G–J Western blot analysis of SEC fractions collected from NIH3T3 AP1/AP2 KO cells stably expressing the short isoform of AP1 (G), long isoform of AP1 (H), short isoform of AP2 (I), and long isoform of AP2 (J). Cell lysates were fractionated on a Superose 6 Increase 3.2/300 column. Circular pictograms below SEC immunoblots schematize PRPS complex configurations. Double circle denotes multiple copies of the protein interacting in a heteromeric complex. Double circle with dotted inner circle denotes multiple copies forming homo-oligomers. Single circle denotes a single protein interacting within the complex. Circle with inner vertical lines denotes proteins forming a trimer or tetramer. Western blot data (A–C, E, G–J) are representative of at least 2 biological repeats. Source data are provided as a Source Data file.
Fig. 6 Translation control of PRPS complex assembly. A–C Western blot analysis of SEC fractions collected from native whole-cell lysates of NIH3T3 AP1 KO cells stably expressing AP1 lacking the non-homologous region (NHR) (A), AP1/AP2 KO cells stably expressing chimeric AP1 with AP2’s N-terminus (residues 1–95) B and chimeric AP2 with AP1’s N-terminus (residues 1–95) (C). D Schematic representation of alternative start sites in mammalian AP1 and AP2 and their consequent translation into short and long isoforms. Base positions correspond to the mouse homologs. TSS1 and TSS2 represent upstream and downstream translation start sites, respectively. E Multiple isoforms for AP1 and AP2 detected under optimal SDS-PAGE resolution. Arrows in immunoblots indicate the longer isoforms of AP1 and AP2, corresponding to N-terminal leader sequences of 29 and 12 amino acids, respectively. F Phylogenetic distribution of PRPS homologs (PRPS1, PRPS2, PRPSAP2, PRPSAP2 with N-terminal leader sequence, PRPSAP1, and PRPSAP1 with N-terminal leader sequence) in chordates. Presence/absence noted across the tree. PRPSAP2 and PRPSAP1 isoforms with N-terminal leader sequences emerged in ancestors of Amniota and Osteichthyes, respectively. G–J Western blot analysis of SEC fractions collected from NIH3T3 AP1/AP2 KO cells stably expressing the short isoform of AP1 (G), long isoform of AP1 (H), short isoform of AP2 (I), and long isoform of AP2 (J). Cell lysates were fractionated on a Superose 6 Increase 3.2/300 column. Circular pictograms below SEC immunoblots schematize PRPS complex configurations. Double circle denotes multiple copies of the protein interacting in a heteromeric complex. Double circle with dotted inner circle denotes multiple copies forming homo-oligomers. Single circle denotes a single protein interacting within the complex. Circle with inner vertical lines denotes proteins forming a trimer or tetramer. Western blot data (A–C, E, G–J) are representative of at least 2 biological repeats. Source data are provided as a Source Data file.CC BY · PubMed Central
Fig. 7 Timeline of PRPS complex evolution in mammals and implications of distinct PRPS assembly states on cellular fitness. A Chronological duplication events of a Class I PRPS encoding gene in indicated eukaryotes. Grey triangle traces the evolutionary trajectory leading to the emergence of mammalian PRPS homologs. Solid vertical lines within the triangle denote the origin of major lineages; dashed lines indicate significant duplication events giving rise to PRPS homologs, with notable functional innovations occurring post-duplication. Class II PRPS, likely present in the LECA, is lost in craniates, followed by rapid expansion of Class I PRPS homologs in jawed vertebrates. Left square bracket indicates a gene duplication event; left round bracket indicates an extension at the N-terminus. Molecular clock-based evolutionary timeline (in millions of years before present, Ma) adapted from ref. 52. B Association of different PRPS complex configurations with cellular fitness in mammals (i) PRPS1 alone undergoes self-assembly to form homo-oligomers. (ii) PRPS1 and AP1 cannot interact directly and form separate homo-oligomers. (iii) PRPS2 binds PRPS1 to form a dimer and disrupts PRPS1 homotypic assemblies. (iv) AP2 preferentially binds with PRPS1 to form a trimer/tetramer to nucleate the complex. (v) AP1 preferentially binds with AP2 and PRPS2 to elongate the PRPS complex via its N-terminus. (vi) The longer isoform of AP1 with flexible N-terminal extension likely caps the complex. The weight of the connectors between the circles representing proteins reflects the predicted strength of protein-protein interactions; heavier lines indicate stronger predicted interactions.
Fig. 7 Timeline of PRPS complex evolution in mammals and implications of distinct PRPS assembly states on cellular fitness. A Chronological duplication events of a Class I PRPS encoding gene in indicated eukaryotes. Grey triangle traces the evolutionary trajectory leading to the emergence of mammalian PRPS homologs. Solid vertical lines within the triangle denote the origin of major lineages; dashed lines indicate significant duplication events giving rise to PRPS homologs, with notable functional innovations occurring post-duplication. Class II PRPS, likely present in the LECA, is lost in craniates, followed by rapid expansion of Class I PRPS homologs in jawed vertebrates. Left square bracket indicates a gene duplication event; left round bracket indicates an extension at the N-terminus. Molecular clock-based evolutionary timeline (in millions of years before present, Ma) adapted from ref. 52. B Association of different PRPS complex configurations with cellular fitness in mammals (i) PRPS1 alone undergoes self-assembly to form homo-oligomers. (ii) PRPS1 and AP1 cannot interact directly and form separate homo-oligomers. (iii) PRPS2 binds PRPS1 to form a dimer and disrupts PRPS1 homotypic assemblies. (iv) AP2 preferentially binds with PRPS1 to form a trimer/tetramer to nucleate the complex. (v) AP1 preferentially binds with AP2 and PRPS2 to elongate the PRPS complex via its N-terminus. (vi) The longer isoform of AP1 with flexible N-terminal extension likely caps the complex. The weight of the connectors between the circles representing proteins reflects the predicted strength of protein-protein interactions; heavier lines indicate stronger predicted interactions.CC BY · PubMed Central

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Metadata

Authors
Karki, B. R., MacMillan, A. C., Vicente-Muñoz, S., Greis, K. D., Romick, L. E., & Cunningham, J. T.
Affiliations
  • University of Cincinnati (US)
  • Cincinnati Children's Hospital Medical Center (US)
Corresponding author
Cunningham, John T.
ORCID
Venue
Nature Communications
Publisher
Springer Science and Business Media LLC
Published
2025-07-08
Volume
16
Issue
1
Pages
6076-6076
ISSN
2041-1723 (ISSN-L)
DOI
10.1038/s41467-025-61216-z
PMC
PMC12238573
OpenAlex
W4412102667
Cited by
11
References
8
Citations by year
2026: 7 · 2025: 4
FWCI
3.31
Type
journal article
Language
en
Open access
Licence
Locations
  • Nature Communications (publishedVersion, cc-by)
  • bioRxiv (Cold Spring Harbor Laboratory) (acceptedVersion, cc-by)
  • PubMed Central (publishedVersion, cc-by)
  • PubMed (publishedVersion)
  • PubMed Central (submittedVersion, cc-by)
  • DOAJ (DOAJ: Directory of Open Access Journals) (submittedVersion, cc-by-sa)
Funders
  • U.S. Department of Health & Human Services | NIH | National Cancer Institute (R01CA230904)
  • U.S. Department of Health & Human Services | NIH | National Institute of General Medical Sciences (R35GM133561)
  • U.S. Department of Health & Human Services | National Institutes of Health (S10OD026717)
Topics
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Biochemical and Molecular Research
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › RNA and protein synthesis mechanisms
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Genomics and Phylogenetic Studies
Keywords
phosphoribosylpyrophosphate synthetase, gene duplication, complex assembly, nucleotide synthesis, eukaryotic evolution, isogenic cell lines, enzyme regulation, mammalian metabolism

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