MIPOL1

The MIPOL1 gene is located at 14q13.3-q21.1 on the plus strand, spanning base pairs 37,197,888 to 37,579,207 (in the human GRCh38 primary assembly, length: 381,320 base pairs), consisting of 15 exons and 11 introns. Some notable genes in its neighborhood include SLC25A21 (mutation of this gene causes synpolydactyly[10]) and FOXA1.

Gene neighborhood of MIPOL1

The unmodified MIPOL1 protein isoform 1 in humans has an isoelectric point of 5.6 and molecular weight 51.5 kDa.[12] Relative to other human proteins, MIPOL1 consists of unusually low amounts of Proline and Glycine and higher amounts of Glutamic acid and Glutamine.[13]

There are at least three known isoforms of this protein in humans produced by alternative splicing: isoform 1, of length 442 amino acids, isoform 2 of length 261 amino acids and isoform 3 of length 169 amino acids.[5]

Fig.1. MIPOL1 domain diagram generated using Prosite MyDomains.[14] The two coiled-coil domains are highlighted in blue. The green line indicates the Nuclear localization signal. Some important phosphorylation sites have been highlighted in red. Studies have shown that phosphorylation is an important modification for controlling nucleus-cytoplasm shuttling, and may therefore play an important role in sub-cellular localization of this protein (by modifying the NLS or NES)[15] O-GlcNACylation site has been highlighted in grey.

Fig.2.Multiple Sequence Alignment of MIPOL1 Orthologs showing conservation of Bipartite Nuclear Localization Signal in Mammalian orthologs (Hsa is human, Lve is Lipotes vexillifer (dolphin), and Mja is Manis javanica (Pangolin)

MIPOL1 contains two coiled-coil domains in its C-terminus at positions 107 – 212 and 253 – 435[5] (shown in Fig.1). A bipartite nuclear localization signal is predicted at position 128 – 143.[16]

Fig.3. Annotated conceptual translation of MIPOL1 isoform 1 showing the most important features. The underlined parts represent the coiled-coil domains. Bolded amino acids are highly conserved even in orthologs as distant as Cnidarians. Other bracketed regions show conserved protein family regions identified such as COG1196 and COG 4372. Exon boundaries are highlighted in blue. Bipartite nuclear localization signal is highlighted in blue. Parts of the protein absent in other isoforms have been highlighted.

The following post-translational modifications are predicted using bioinformatics tools for MIPOL1.[17] Multiple phosphorylation sites are predicted for this protein, that are conserved in close orthologs, including a Casein kinase 1 (CK1) site, three Casein kinase 2 (CK2) sites, and three NEK2 sites.[18]

Fig.4.Tertiary structure of MIPOL1 generated using I-TASSER:[25] Blue represents the N-terminus, Red represents the C-terminus

The exact structure of the MIPOL1 has not yet been characterized. Homology-based and de novo predictions of its tertiary structure suggest that it may consist of inter-twined alpha helices, forming coiled-coil domains (see Fig.4.).[26][25]

Immunofluorescence imaging in the human U2OS cell line (bone Osteosarcoma epithelial cells) shows localization in the cytosol.[27] Immunohistochemistry imaging of human prostate tissue also suggests cytosolic localization.[28] A bipartite nuclear localization signal is predicted at position 128 – 143, which is highly conserved in mammalian orthologs (see Fig.2.), indicating possible localization in the nucleus.[16]

This protein is known to interact with multiple human proteins, verified via two-hybrid screening. A few notable examples include:

LATS2: Negatively regulates YAP1 in the Hippo signaling pathway that plays a pivotal role in organ size control and tumor suppression by restricting cell proliferation and promoting apoptosis.[39]

ZGPAT (Zinc finger CCCH-type with G patch domain-containing protein): A transcription repressor that negatively regulates expression of EGFR, a gene involved in cell proliferation, survival and migration, suggesting that it may act as a tumor suppressor.[40]

RCOR3 (REST Corepressor 3): A protein that may act as a component of a co-repressor complex that represses transcription[41]

It also interacts with viral proteins such as:

Replicase polyprotein 1ab (SARS-CoV2): A multifunctional protein involved in the transcription and replication of viral RNAs.[38]

Protein E7 (Human Papillomavirus): Plays a role in viral genome replication by driving entry of quiescent cells into the cell cycle.[42]

The MIPOL1 protein has no known paralogs in humans and other species for which orthologs have been found, therefore, it is the only member of its gene family.

There are more than 300 known orthologs of the MIPOL1 protein in Animalia, ranging from primates to corals and sea anemones in phylum Cnidaria.[43] Orthologs of the protein were found in species as distant as Trichoplax adhaerens, a simple primitive invertebrate species. Table 2 shows a sample of the ortholog space.

Closely related orthologs are found in chordates such as mammals, reptiles, birds and amphibians, with sequence similarities greater than 70%. Sequence lengths of orthologs were similar to the human MIPOL1 protein, with no significant gene duplication observed.

Organisms with sequence similarities in the 55-70% range (moderately related orthologs) were found in bony fish, cartilaginous fish and coelacanths. Sequence length is generally longer in these species, with a longer amino acid sequence in the N-terminus (alignment with human protein occurs around amino acid 100).

Distantly related orthologs with similarities less than 50% (around 30 – 40%) are found in hemichordates, echinoderms, arthropods, molluscs, cnidaria and placozoa. Multiple sequence alignment with distant orthologs indicates poor alignment in the N-terminus of the protein.

Two COG (Clusters of Orthologous Groups of proteins) domains were found in this protein (see Fig.3): COG1196 at position 106 - 340 (Chromosome segregation ATPase[44]) and COG4372 at 259 - 431 (uncharacterized conserved protein containing a DUF3084 domain[45])[46]

Fig.5. Plot of number of amino acid changes per 100 amino acids as a function of date of divergence for MIPOL1, cytochrome c and Fibrinogen alpha

Using a linear regression analysis on a plot of corrected percent divergence (amino acid changes per 100 amino acids) as a function of date of divergence from humans for different MIPOL1 orthologs (see Fig.5), it is estimated that a 1% change in amino acids in the MIPOL1 protein takes 5.68 million years. MIPOL1 protein is evolving at a moderate rate relative to fast evolving protein such as fibrinogen alpha, and slow evolving proteins such as cytochrome C.