Dopaminylation

Protein monoaminylation was first identified in 1957 by Heinrich Waelsch and colleagues at Columbia University. After discovering that primary amines could be covalently incorporated into proteins via transamidation at glutamine residues, Despite its discovery in the mid-twentieth century, monoaminylation was not investigated as a post-translational modification until 2003, when Diego Walther and colleagues at the Max-Planck-Institute for Molecular Genetics revealed that serotonylation of small GTPases mediates ⍺-granule release during the activation and aggregation of platelets. Notably, monoaminylation was not uncovered as an epigenetic regulatory mechanism until 2019, when Lorna Farrelly and colleagues at the Icahn School of Medicine reported the H3Q5-serotonylation (H3Q5ser) modification for the first time. Later, in 2020, the H3Q5-dopaminylation (H3Q5dop) modification was identified in the striatum by Ashley Lepack and colleagues also at the Icahn School of Medicine. Five years later, Qingfei Zheng and colleagues at Ohio State University discovered the H3Q5-histaminylation (H3Q5his) modification in the posterior hypothalamic tuberomammillary nucleus (TMN). In 2024, Nan Zhang and colleagues at Ohio State University developed a bicyclononyne (BCN)-containing probe to address the lack of efficient pan-specific antibodies targeting dopaminylated glutamine. Herein, the group successfully applied the BCN-probe in chemical proteomic profiling of the dopaminylation proteome in a colorectal cancer cell line (ie., HCT 116 cells), identifying over 400 dopaminylated proteins and providing extensive pathway enrichment data following their analyses using the Kyoto Encyclopedia of Genes and Genomes (KEGG) database. == Mechanism ==

Dopaminylation is catalyzed by transglutaminase 2 (TGM2) in a calcium-dependent manner, and relies upon the intracellular bioavailability of dopamine substrates. Generally, protein dopaminylation occurs in the cytoplasm; however, histone dopaminylation only occurs within the nucleus. The active site itself is composed of a well conserved catalytic triad (Cys277–His335–Asp358) situated within a substrate binding channel, which is bordered by two conserved residues (Trp241 and Trp332) that facilitate catalysis through stabilization of the transition state. Once intracellular Ca2+ binds to TGM2 and exposes the substrate binding channel, the glutamine residue of a substrate protein (i.e., histone H3, TPD1) is free to enter the enzyme active site. As a transamidation reaction, the mechanism for protein dopaminylation can be summarized in two parts: an initial thioester formation, followed by isopeptide bond formation. Fig. 1 Mechanism for Protein Dopaminylation Dopaminylation is a two step, Ca2+-dependent reaction in which TGM2 catalyzes the covalent attachment of dopamine onto the glutamine residue of a substrate protein (i.e., Histone H3, TPI1). (A) The catalytic cysteine residue (Cys277) of TGM2 facilitates an initial acyl transfer reaction, which is ultimately followed by isopeptide bond formation (B). When intracellular Ca2+ and dopamine concentrations are sufficient, TGM2-catalyzed dopaminylation of substrate proteins can occur. First, the catalytic cysteine residue (Cys277) in the TGM2 active site nucleophilically attacks the 𝛾-carboxamido group of the glutamine residue in an acyl transfer reaction (Fig. 1A), forming a thioester intermediate and releasing one molecule of ammonia (NH3) as a result. Next, the deprotonated primary amine of the dopamine substrate nucleophilically attacks the 𝛾-thioester group of the intermediate, forming a stable isopeptide bond and ultimately releasing the enzyme (Fig. 1B). == Functions ==

Histone Dopaminylation With the discovery of histone monoaminylation in 2019, monoaminylation thus entered into the complex and ever-growing field of epigenetics, posing as a dynamic set of novel regulatory mechanisms. Data as to the effects of H3Q5dop are displayed in detail within the table below (Table 1). Monoaminylation Tissue (or Cell) Type Modification Biological Function References Dopaminylation Ventral Tegmental Area (VTA) H3Q5dopH3K4me3Q5dop Promotes relapse-like behavior and modulates neuronal gene expression patterns in the VTA following cocaine consumption (Lepack et al., 2020) Herein, failure to recognize novel odor was reportedly linked to increased dopamine transmission, decreased levels of TGM2, and increased histone trimethylation (H3K4me3) and dopaminylation (H3Q5dop) in the amygdala following exposure to early-life stressful social experience (SSE). It remains unclear whether the reported fluctuations in TGM2 levels could be attributed to changes in TGM2 expression levels or changes in TGM2 activity levels. Ambiguity aside, this data provides useful insight, as early-life adversity paradigms appear sufficient for reconfiguration of epigenetic signatures within the limbic system, thereby establishing stable, differential epigenetic programs which may contribute to lifelong susceptibility for affective psychopathologies (i.e., major depressive disorder, bipolar disorder, anxiety disorders). Protein Dopaminylation While research has demonstrated several roles for histone dopaminylation in modulating transcriptional and behavioral plasticity, the cellular dopaminylated proteome has remained poorly understood, largely in part due to the lack of efficient pan-specific antibodies targeting dopaminylated glutamine. However, a recently developed bicyclononyne (BCN)-containing probe has been successfully applied in chemical proteomic profiling of the dopaminylation proteome in cancer cells. Herein, authors present emerging evidence suggesting that 425 proteins possessed dopaminylation sites in a colorectal cancer cell line (ie., HCT 116 cells). Further analysis using the Kyoto Encyclopedia of Genes and Genomes (KEGG) database revealed that these dopaminylated proteins were involved in numerous signal transduction pathways, RNA processing pathways, and several disease-associated signaling pathways. Details as to the identified dopaminylated proteins are displayed in the tables below. However, only 176 of the 425 identified dopaminylated proteins were named by the authors within their supplementary materials, and as such, only those 176 proteins can be discussed within this article. Nevertheless, to benefit this article, the list of 176 proteins has been reanalyzed using the KEGG database, and the details of the identified KEGG PATHWAY and KEGG BRITE search results are displayed in the data tables below. The following data table lists the dopaminylated proteins (ie., out of those 176 proteins) identified by KEGG BRITE across 10 protein families related to genetic information processing: BRITE Category Identified Dopaminylated Proteins Reference Spliceosome ALYREFCBX3FRG1FUBP1FUSHNRNPA2B1HNRNPA3HNRNPDLHNRNPFHNRNPH1HNRNPH3HNRNPKHNRNPLLLUC7LMBNL1NSRP1PQBP1PRPF8PSPC1RBM15BRBM22RBM4RBMXL1RP9SAFBSAFB2SCAF4SDE2SF3A3SFPQSMNDC1SNRPASNRPCSONSRRM2SRSF11SUGP1TCERG1WBP11ZNF326 (Zhang et al., 2024) mRNA Biogenesis ALYREFAPOBEC3BAPOBEC3FCASC3CNOT2CPSF2CPSF4FIP1L1FXR2HNRNPA2B1LIMD1NUP133NUP50NUP58NUP62NUP98PCF11RBM15BSUPT5HTAF15TNRC6ATNRC6BZC3H14ZFP36L2 (Zhang et al., 2024) Transcription Machinery ARID2CPSF4HEXIM1IWS1MED1MED12MED23PCF11POLR2BSCAF4SFPQSUPT5HTAF15TAF6LZC3H8ZNF143ZNF326 (Zhang et al., 2024) Chromosome & Associated Proteins AEBP2ANKRD17ARID2ARID4BBPTFBRCA1CARM1CBX3CBX4CDCA5CENPFCLSPNDNTTIP2DRG1ESCO1FLOT1HAT1HMGB3ING1JMJD1CLIMD1NCAPG2NCOA3NCOR1NPM1NSD1NUMA1NUP133NUP98PAXIP1RBBP5RESTSTAG2TAF6LTELO2TNRC6ATNRC6BUBN2YY1ZMYM3ZNF217 (Zhang et al., 2024) Ribosome Biogenesis DNTTIP2HEATR1NIFKNOL9NPLOC4PWP1PWP2TBL3WDR75 (Zhang et al., 2024) Ribosome Biogenesis DNTTIP2HEATR1NIFKNOL9NPLOC4PWP1PWP2TBL3WDR75 (Zhang et al., 2024) DNA Repair & Recombination Proteins BRCA1CLSPNDUTPOLD1POLR2BSDE2TELO2UIMC1 (Zhang et al., 2024) DNA Replication Proteins POLD1SDE2TELO2 (Zhang et al., 2024) tRNA Biogenesis DARS2ELAC2ELP2 (Zhang et al., 2024) Translation Factors DRG1EEF1D (Zhang et al., 2024) The KEGG database is composed of multiple database modalities, in which data on various proteins, biological pathways, and diseases are listed for bioinformatics research. Enrichment analysis of gene-sets is typically performed using the web-based tool ShinyGO, within which KEGG databases may be used to convert lists of genes into details on functional hierarchies and pathway enrichment. Notably, KEGG PATHWAY is a collection of manually annotated pathway maps, which illustrate various enzyme-catalyzed reactions and detail their components. Moreover, KEGG BRITE is another database listed within the KEGG program, which acts as a functional hierarchy viewer and allows users to identify higher-order cellular functions from large-scale datasets. With this in mind, the following data table lists the dopaminylated proteins (ie., out of those 176 proteins) identified by KEGG BRITE across 14 protein families related to genetic information processing or signaling and cellular processes: BRITE Category Identified Dopaminylated Proteins Reference Enzymes ABL1BRCA1CARM1DARS2DUTELAC2HAT1MTORNCOA3NSD1OTUD4POLD1POLR2BPRDX1PRDX5PRKCDRNF138TAOK1USP47USP7 (Zhang et al., 2024) Transcription Factors AEBP2ARID2ARID4BCICCNBPCREB1FOXJ3GLI2KLF5LRRFIP1NFX1RAI1RERERESTSP1SP3STAT1TFCP2YY1ZBTB7AZC3H8ZFXZHX1ZHX3ZNF16ZNF638ZNF668ZNF8ZSCAN26 (Zhang et al., 2024) Protein Phosphatases & Associated Proteins AXIN1BRCA1MAPTNCOR1SFPQTSC2UBN2WBP11YLPM1 (Zhang et al., 2024) Ubiquitin System BRCA1CBX4NFX1RNF138USP47USP7ZBTB7A (Zhang et al., 2024) Membrane Trafficking BAG3FLOT1MAPTMTORNCOA7 (Zhang et al., 2024) Cytoskeleton Proteins FHOD1LMNAMAPTTMPO (Zhang et al., 2024) Protein Kinases ABL1MTORPRKCDTAOK1 (Zhang et al., 2024) Domain-containing Proteins Not Elsewhere Classified FHL2LRWD1PXNSORBS3 (Zhang et al., 2024) Peptidases & Inhibitors OTUD4USP7USP47 (Zhang et al., 2024) Glycosaminoglycan Binding Proteins C1QBPHDGF (Zhang et al., 2024) Chaperones & Folding Catalysts BAG3CALR (Zhang et al., 2024) Proteasome PSMD4PSMD10 (Zhang et al., 2024) Amino Acid-Related Enzymes DARS2 (Zhang et al., 2024) Lectins CALR (Zhang et al., 2024) KEGG pathway maps are an invaluable resource for visualizing enzyme-catalyzed reactions and the details of their components. Pathway maps have been generated for a vast number of human diseases, from cancers (ie., by type) to substance addiction (ie., by drug) to neurodegenerative and prion diseases. The following data table lists the dopaminylated proteins (ie., out of those 176 proteins) identified by KEGG PATHWAY across a number of human diseases, with the link to each disease pathway map provided below: Disease or Infection KEGG Pathway Map Link Identified Dopaminylated Proteins Reference Amyotrophic Lateral Sclerosis KEGG ALS Pathway Map ALYREFNUP50DAXXHNRNPA3NUP62MTORFUSHNRNPA2B1NUP98NUP133PSMD4NUP58 (Zhang et al., 2024) Transcriptional Misregulation in Cancer KEGG Transcriptional Misregulation in Cancer Map JMJD1CFUSSP1TAF15NCOR1 (Zhang et al., 2024) Breast Cancer KEGG Breast Cancer Pathway Map MTORSP1BRCA1NCOA3AXIN1 (Zhang et al., 2024) HIV-1 Infection KEGG HIV-1 Infection Pathway Map APOBEC3FAPOBEC3BCALRMTORPXNTAB2 (Zhang et al., 2024) Huntington's Disease KEGG Huntington's Disease Pathway Map CREB1MTORPOLR2BPSMD4RESTSP1 (Zhang et al., 2024) Chemical Carcinogenesis KEGG Chemical Carcinogenesis Pathway Map (Receptor Activation)KEGG Chemical Carcinogenesis Pathway Map (ROS) ABL1CREB1KLF5MTORPRKCD (Zhang et al., 2024) HPV Infection KEGG HPV Infection Pathway Map AXIN1CREB1MTORNFX1PXNSTAT1TSC2 (Zhang et al., 2024) General Pathways in Cancer KEGG General Pathways in Cancer Map ABL1AXIN1GLI2MTORNCOA3SMAD4SP1STAT1 (Zhang et al., 2024) HSV-1 Infection KEGG HSV-1 Infection Pathway Map ALYREFCALRDAXXMTORSTAT1TAB2TSC2 (Zhang et al., 2024) Shigellosis KEGG Shigellosis Pathway Map CBX3MTORPRKCDPXNTAB2TAB3 (Zhang et al., 2024) Alzheimer's Disease KEGG Alzheimer's Disease Pathway Map AXIN1MAPTMTORPSMD4 (Zhang et al., 2024) Parkinson Disease KEGG Parkinson Disease Pathway Map DAXXMAPTPSMD4 (Zhang et al., 2024) MicroRNAs in Cancer KEGG MicroRNAs in Cancer Pathway Map CDCA5MTORABL1HNRNPKBRCA1 (Zhang et al., 2024) Spinocerebellar Ataxia/td> KEGG Spinocerebellar Ataxia Pathway Map CICMTORPSM4PUM2SP1 (Zhang et al., 2024) Cushing Syndrome/td> KEGG Cushing Syndrome Pathway Map AXIN1CREB1RBBP5SP1 (Zhang et al., 2024) Hepatocellular Carcinoma KEGG Hepatocellular Carcinoma Pathway Map ARID2MTORSMAD4AXIN1 (Zhang et al., 2024) Viral Carcinogenesis KEGG Viral Carcinogenesis Pathway Map CREB1HNRNPKPXNUSP7 (Zhang et al., 2024) Gastric Cancer KEGG Gastric Cancer Pathway Map AXIN1MTORSMAD4 (Zhang et al., 2024) Pancreatic Cancer KEGG Pancreatic Cancer Pathway Map MTORSMAD4STAT1 (Zhang et al., 2024) Colorectal Cancer KEGG Colorectal Cancer Pathway Map AXIN1MTORSMAD4 (Zhang et al., 2024) Choline Metabolism in Cancer KEGG Choline Metabolism in Cancer Pathway Map MTORSP1TSC2 (Zhang et al., 2024) Basal Cell Carcinoma KEGG Basal Cell Carcinoma Pathway Map AXIN1GLI2 (Zhang et al., 2024) Chronic Myeloid Leukemia KEGG Chronic Myeloid Leukemia Pathway Map ABL1SMAD4 (Zhang et al., 2024) Neutrophil Extracellular Trap (NET) Formation KEGG NET Pathway Map HAT1MTOR (Zhang et al., 2024) Prostate Cancer KEGG Prostate Cancer Pathway Map CREB1MTOR (Zhang et al., 2024) Glioma KEGG Glioma Pathway Map MTOR (Zhang et al., 2024) Endometrial Cancer [... KEGG Endometrial Cancer Pathway Map] AXIN1 (Zhang et al., 2024) KEGG pathway maps are also organized for individual signal transduction pathways (ie., Hippo signaling, JAK-STAT pathway, etc). Pathway maps have been generated for a vast number of cell signaling cascades, from canonical pathways (ie., Wnt signaling) to broader endocrine systems (ie., estrogen signaling) to more specified signaling molecules and interactions (ie., IgSF CAM signaling). The following data table lists the dopaminylated proteins (ie., out of those 176 proteins) identified by KEGG PATHWAY across a number of signal transduction pathways, with the link to each signaling pathway map provided in the table below: Signaling Pathway KEGG Pathway Link Identified Dopaminylated Proteins Reference AMPK Signaling Pathway KEGG AMPK Signaling Pathway Map CREB1MTORTSC2 (Zhang et al., 2024) Apelin Signaling Pathway KEGG Apelin Signaling Pathway Map MTORSMAD4 (Zhang et al., 2024) C-Type Lectin Receptor SignalingPathway KEGG C-Type Lectin Receptor Signaling Pathway Map PRKCDSTAT1 (Zhang et al., 2024) Cadherin Signaling KEGG Cadherin Signaling Pathway Map ATN1AXIN1ERBINLIMD1RERETAOK1 (Zhang et al., 2024) Chemokine Signaling Pathway KEGG Chemokine Signaling Pathway Map PRKCDPXNSTAT1 (Zhang et al., 2024) ErbB Signaling Pathway KEGG ErbB Signaling Pathway Map ABL1MTOR (Zhang et al., 2024) Estrogen Signaling Pathway KEGG Estrogen Signaling Pathway Map CREB1NCOA3PRKCDSP1 (Zhang et al., 2024) Hippo Signaling Pathway KEGG Hippo Signaling Pathway Map LIMD1 (Zhang et al., 2024) IgSF CAM Signaling KEGG IgSF CAM Signaling Pathway Map ABL1PXN (Zhang et al., 2024) Insulin Signaling Pathway KEGG Insulin Signaling Pathway Map FLOT1MTORTSC2 (Zhang et al., 2024) IL-17 Signaling Pathway KEGG IL-17 Signaling Pathway Map TAB2TAB3 (Zhang et al., 2024) Integrin Signaling Pathway KEGG Integrin Signaling Pathway Map MTORPXN (Zhang et al., 2024) JAK-STAT Signaling Pathway KEGG JAK-STAT Signaling Pathway Map MTORSTAT1 (Zhang et al., 2024) MAPK Signaling Pathway KEGG MAPK Signaling Pathway Map DAXXMAPTTAB2TAOK1 (Zhang et al., 2024) mTOR Signaling Pathway KEGG mTOR Signaling Pathway Map MTORTELO2TSC2 (Zhang et al., 2024) Neurotrophin Signaling Pathway KEGG Neurotrophin Signaling Pathway Map ABL1PRKCD (Zhang et al., 2024) NF-kappa B Signaling Pathway KEGG NF-kappa B Signaling Pathway Map TAB2TAB3 (Zhang et al., 2024) NOD-like Receptor Signaling Pathway KEGG NOD-like Receptor Signaling Pathway Map ERBINPRKCDSTAT1TAB2TAB3 (Zhang et al., 2024) Phospholipase D Signaling Pathway KEGG Phospholipase D Signaling Pathway Map MTORTSC2 (Zhang et al., 2024) PI3K-Akt Signaling Pathway KEGG PI3K-Akt Signaling Pathway Map BRCA1CREB1MTORTSC2 (Zhang et al., 2024) TGF-beta Signaling Pathway KEGG TGF-beta Signaling Pathway Map NCOR1SMAD4SP1 (Zhang et al., 2024) Thyroid Hormone Signaling Pathway KEGG Thyroid Hormone Signaling Pathway Map MED1MED12MTORNCOA3NCOR1STAT1TSC2 (Zhang et al., 2024) Toll-Like Receptor Signaling Pathway KEGG Toll-Like Receptor Signaling Pathway Map STAT1TAB2 (Zhang et al., 2024) TNF Signaling Pathway KEGG TNF Signaling Pathway Map CREB1TAB2TAB3 (Zhang et al., 2024) Wnt Signaling Pathway KEGG Wnt Signaling Pathway Map AXIN1SMAD4 (Zhang et al., 2024) == See Also ==