Evolution and function of calponin and transgelin

Abstract

Calponin and transgelin (originally named SM22) are homologous cytoskeleton proteins that regulate actin-activated myosin motor functions in smooth muscle contraction and non-muscle cell motility during adhesion, migration, proliferation, phagocytosis, wound healing, and inflammatory responses. They are abundant cytoskeleton proteins present in multiple cell types whereas their physiological functions remain to be fully established. This focused review summarizes the evolution of genes encoding calponin and transgelin and their isoforms and discusses the structural similarity and divergence in vertebrate and invertebrate species in the context of functions in regulating cell motility. As the first literature review focusing on the evolution of the calponin-transgelin family of proteins in relevance to their structure-function relationship, the goal is to outline a foundation of current knowledge for continued investigations to understand the biological functions of calponin and transgelin in various cell types during physiological and pathological processes.

Keywords: calponin; cell motility; cytoskeleton; evolution; isoform genes; structure-function relationship; transgelin.

FIGURE 1

Phylogenetic tree of vertebrate calponin and isoforms. The phylogenetic tree was generated by alignment of amino acid sequences of calponin isoforms in representative vertebrate species with the DNASTAR MegAlign computer software (Lasergene, lnc, Madison, WI) using the Clustal W method. The sequence similarity-derived evolutionary lineages demonstrate that each of the calponin isoforms is conserved in the vertebrate phylum while the three isoforms have significantly diverged during vertebrate evolution. This pattern indicates the adaptation of the calponin isoforms to different cell functions and tissue environments. Calponin isoforms 1, 2, and 3 are marked in purple, red and green, respectively. The degrees of evolutionary divergence are indicated by the lengths of lineage lines. The NCBI database accession numbers of the sequences analyzed are: African clawed frog CNN1, NP_001085014.1; African clawed frog CNN2, ABG49504.1; African clawed frog CNN3, NP_001080482.1; Atlantic salmon CNN1, NP_001139857.1; Atlantic salmon CNN2, NP_001133873.1; Atlantic salmon CNN3, NP_001133337.1; Band tailed pigeon CNN2, OPJ86593.1; Bighead croaker CNN1, TKS92442.1; Bighead croaker CNN2, TKS80042.1; Bighead croaker CNN3, TKS88103.1; Brown tree snake CNN2, JAG68493.1; Chimpanzee CNN1,NP_001267033.1; Chimpanzee CNN2, JAA13388.1; Chimpanzee CNN3, JAA44470.1; Chinese hamster CNN1, EGV96164.1; Chinese hamster CNN2, EGV99480.1; Chinese hamster CNN 3, EGW11625.1; Chicken CNN1, NP_990847.1; Chicken CNN2, NP_001135728.1; Chicken CNN3, NP_001341600.1; Copperhead snake CNN2, JAV51035.1; Eastern Diamondback rattlesnake CNN2, AFJ49586.1; Human CNN1, NP_001290.2; Human CNN2, AAI48265.1; Human CNN3, AAB35752.1; Mainland tiger snake CNN1, XP_026535408.1; Mouse CNN1, AAI38864.1; Mouse CNN2, EDL31614.1; Mouse CNN3, AAH85268.1; Naked mole CNN1, JAN96391.1; Naked mole CNN2, EHB16944.1; Naked mole CNN3, XP_004841280.1; Snakehead fish CNN1, KAF3700611.1; Snakehead fish CNN2, KAF3699176.1; Snakehead fish CNN3, KAF3703402.1; Western terrestrial garter snake CNN1, XP_032066879.1; Western clawed frog CNN1, NP_001015796.1; Western clawed frog CNN2, NP_998841.1; Western clawed frog CNN3, NP_989257.1; Western terrestrial garter snake CNN2, XP_032064388.1; Western terrestrial garter snake CNN3, XP_032074010.1; Zebrafish CNN1, XP_701038.5; Zebrafish CNN2, NP_998514.1; Zebrafish CNN3, NP_956047.1.

FIGURE 2

Phylogenetic tree of vertebrate transgelin isoforms. The phylogenetic tree was generated by aligning amino acid sequences of the three transgelin isoforms in representative vertebrate classes including fish, amphibian, reptile, avian, and mammal with the MegAlign computer program (Lasergene; DNASTAR, lnc, Madison, WI) using Clustal W method. The degrees of evolutionary divergence are indicated by the lengths of lineage lines. TAGLN, TAGLN2, and TAGLN3 isoforms are marked in purple, red and green, respectively. The NCBI database accession numbers of the sequences analyzed are: African clawed frog TAGLN, NP_001083600.1; African clawed frog TAGLN2 NP_001080783.1; African turquoise killifish TAGLN3, KAF7213900.1; Black-legged kittiwake TAGLN, XP_054078918.1; Black-legged kittiwake TAGLN2, XP_054038817.1; Black-legged kittiwake TAGLN3, XP_054071416.1; Cape cliff lizard TAGLN, XP_053124973.1; Cape cliff lizard TAGLN2, XP_053133737.1; Cape cliff lizard TAGLN3, XP_053167629.1; Chicken TAGLN, AAA48782.1; Chicken TAGLN2, XP_024999416.1; Chicken TAGLN3, XP_040518015.1; Diamondback rattlesnake TAGLN, JAI10881.1; European plaice TAGLN, XP_053298355.1; European seabass TAGLN, XP_051257332.1; Green swordtail TAGLN3, KAF5894368.1; Human TAGLN, NP_001001522.1; Human TAGLN2, KAI4083433.1; Human TAGLN3, KAI2530827.1; Mexican tetra TAGLN2, XP_049332440.1; Mexican tetra TAGLN3, KAG9283353.1; Mouse TAGLN, CAA92941.1; Mouse TAGLN2, NP_848713.1; Mouse TAGLN3, AAH55338.1; Orange spotted grouper TAGLN, ABW04145.1; Tiger rattlesnake TAGLN, 2, XP_039222054.1; Tiger rattlesnake TAGLN3, XP_039220015.1; Tiny caecilian TAGLN, XP_030076731.1; Tiny caecilian TAGLN2, XP_030043052.1; Tiny caecilian TAGLN3, XP_030060064.1; Two-lined caecilian TAGLN, XP_029429188.1; Two-lined caecilian TAGLN2, XP_XP_029436331.1; Two-lined caecilian TAGLN3, XP_029434652.1; Western clawed frog TAGLN, NP_001025579.1; Western clawed frog TAGLN2, NP_989354.1; Yellow drum fish TAGLN, KAG8008337.1.

FIGURE 3

Phylogenetic tree of invertebrate calponin isoforms. The phylogenetic tree of invertebrate calponin was generated by aligning the amino acid sequences of annotated calponin isoforms in the representative invertebrate species with the MegAlign computer program (Lasergene; DNASTAR, lnc, Madison, WI) using Clustal W method. The degrees of evolutionary divergence are indicated by the lengths of lineage lines. Calponin isoforms 1, 2, and 3 are indicated in purple, red and green fonts, respectively. The NCBI database accession numbers of the sequences analyzed are: Bark spider CNN2, GIY95920.1; Bat sea star CNN1, XP_038068192.1; Bat sea star CNN3, XP_038068193.1; Belted springtail CNN1, ODN05277.1; Bird tapeworm CNN3, JAP59453.1; Blood fluke CNN3, XP_012795279.2; Dog round worm CNN2, KHN73181.1; Dog tapeworm CNN1, KAH9282893.1; Dog tapeworm CNN3, EUB56891.1; East Asian octopus CNN1, XP_029648512.1; Fungivorous round worm CNN2, KAH7720906.1; Garden spider CNN3, GBN55272.1; Gray mussel CNN2, APB61452.1; Hood coral CNN1, PFX34187.1; Hood coral CNN3, PFX34186.1; Human whip worm CNN1, CDW60276.1; Intoshia linei worm CNN1, OAF71457.1; Japanese scallop CNN3, OWF41392.1; Oriental liver fluke CNN3, GAA43144.2; Pale anemone CNN1,KXJ22199.1; Taxon placozoa CNN1, RDD43626.1; Water flea CNN3, JAM94970.1; White springtail CNN1, OXA37663.1; Satsuma tube worm CNN3, KAI0207068.1; Salmon louse CNN3, ACO12641.1; Sea louse CNN3, ACO14747.1; Sea spider CNN3, KAG1690965.1; Spider mite CNN3, XP_015791637.1.

FIGURE 4

Phylogenetic tree of invertebrate animal and fungal transgelin isoforms. The phylogenetic tree was generated by aligning amino acid sequences of transgelin isoforms of representative invertebrate species with the DNASTAR MegAlign computer software (Lasergene, lnc, Madison, WI) using Clustal W method. Transgelin isoforms 1, 2, and 3 are listed in purple, red and green fonts, respectively. Fungal transgelins are highlighted in brown font. The NCBI database accession numbers of the sequences are: California two spot octopus TAGLN3, XP_014777580.1; Caenorhabditis elegans TAGLN, NP_493713.2; Common house spider TAGLN, LAA01862.1; Common starfish TAGLN, QAA95982.1; Darwini’s bark spider TAGLN3, GIY80101.1; Dog tapeworm TAGLN, KAH9282095.1; Oriental liver fluke TAGLN, KAG5444608.1; Freshwater snail TAGLN3, KAH9519225.1; Hood coral TAGLN2, PFX34185.1; Human scabies TAGLN2, KAI7688587.1; Japanese scallop TAGLN3, OWF50751.1; Mediterranean mussel TAGLN, VDI10566.1; Oriental lung fluke TAGLN, KAA3679206.1; Pharaoh cuttlefish TAGLN, CAE1295289.1; Pork worm TAGLN2, KRY33709.1; Satsuma tube worm TAGLN2, KAI0231824.1; Sea slug TAGLN, GFO13790.1; Silkworm moth TAGLN, ABF51271.1; Speckled wood butterfly TAGLN, JAA92459.1; Strawberry crimp disease nematode TAGLN, KAI6193121.1; Water flea TAGLN3, JAJ10604.1; Aspergillus luchuensis TAGLN, GAT26957.1; Budding yeast TAGLN, KAF4004496.1; Candida parapsilosis TAGLN, KAI5906036.1; Fission yeast TAGLN, O14185.1.

FIGURE 5

Cytoskeleton and regulatory partners of calponin and transgelin. (A) Multiple cytoskeleton and regulatory proteins have been identified to interact with calponin. Calponin binds actin filaments through two actin-binding sites (ABS), tropomyosin via the N-terminal calponin homology (CH) domain, and gelsolin via the actin binding sites. Calponin interacts with microtubules through the actin-binding sites and the repeating motifs whereas with desmin through the region of CH domain and the actin-binding sites. Residues 144–182 of calponin interact with myosin. Ca²⁺-calmodulin and Ca²⁺-S100 bind calponin at the actin-binding sites, which releases the inhibitory effect on myosin MgATPase. An N-terminal fragment of calponin was reported to interact with phospholipids and the CH domain overlaps with an ERK binding region. (B) In transgelin, the CH domain binds ERK, the ABS domain binds F-actin, and the C-terminal calponin like (CLIK) segment binds ezrin. Transgelin interacts with ezrin through its CLIK domain. Two EF-hand Ca binding motifs are found in the CH domain of transgelin, suggesting a possible regulation via calcium signaling.

FIGURE 6

Structural comparison of vertebrate calponin and transgelin isoforms. (A) The primary structural comparison of three calponin isoforms shows the N-terminal calponin homology (CH) domain, the two actin-binding sites (ABS), the three calponin like repeat (CLIK) motifs, and the C-terminal variable region that constitutes the main differences between the isoforms. Conserved PKC phosphorylation sites in all three isoforms include Ser₁₇₅ and Thr₁₈₄ (blue arrow) in the second actin-binding site overlapping with the first CLIK motif. Calponin 1 has two tyrosine phosphorylation sites at Tyr₂₆₁ and Tyr₁₈₂ (black arrow). Calponin 2 has a potentially phosphorylation site at Ser₁₇₇ (green arrow). Calponin 3 has two ROCK phosphorylation sites at Ser₂₉₃ and Ser₂₉₆ as well as an MEKK1 phosphorylation site at Thr₂₈₈ in the C-terminal region. (B) The structure of human transgelin consists of three regions: An N-terminal CH domain, an ABS, and a C-terminal CLIK region, which resembling a mini-calponin lacking the C-terminal variable tail. Phosphorylation sites have been found in transgelin-2 at Ser₁₁, Ser83, Thr₈₄, Ser₁₄₅, Ser₁₆₃, Ser₁₈₀, Ser₁₈₅, Thr₁₉₀, and Tyr₁₉₂. In transgelin-3, phosphorylation sites have been identified at Ser₁₁, Thr₁₂₈, Ser₁₃₄, Ser₁₆₃, Ser₁₈₀, Ser₁₈₅, and Thr₁₉₀, and a methylation site at Arg₁₉₆.

FIGURE 7

Linear structural maps of invertebrate calponin-like and transgelin-like proteins. The structures of invertebrate calponin-transgelin family proteins are compared. The blue box indicates the N-terminal calponin homology (CH) domain and the red ellipses are calponin like (CLIK) motifs. The invertebrate calponin-transgelin family proteins contain a CH domain and highly diverse numbers of CLIK repeats whereas the calponin-related protein unc-87 of Caenorhabditis elegans does not have the CH domain (Ono, 2021).

FIGURE 8

Phylogeny of the calponin-transgelin superfamily genes in vertebrates. The combined phylogenetic tree of vertebrate calponin and transgelin isoforms was generated by aligning amino acid sequences of representative species with the DNASTAR MegAlign computer software (Lasergene, lnc, Madison, WI) using Clustal W method. Transgelin isoforms 1, 2, and 3 are marked in purple, red and green fonts, respectively. The vertebrate calponin isoforms 1, 2, and 3 are clustered in the blue box. The phylogenetic lineages demonstrate that calponin and transgelin diverged early during vertebrate evolution and transgelin-1 emerged before transgelin-2, transgelin-3 and the three calponin isoforms. The NCBI database accession numbers of the sequences are listed in the legends of Figures 1, 2.

FIGURE 9

Overall phylogeny of calponin-transgelin genes in vertebrate and invertebrate animals. The combined phylogenetic tree of vertebrate and invertebrate calponin and transgelin was generated by aligning amino acid sequences of representative species with the DNASTAR MegAlign computer software (Lasergene, lnc, Madison, WI) using Clustal W method. Vertebrate calponin isoforms are in blue font, invertebrate calponins are in black font, vertebrate transgelins are in green font and invertebrate transgelins are in red font. The NCBI database accession numbers of the sequences are in the legends of Figures 1–4. The evolutionary lineages show that while calponin and transgelin have evolved into distinct and conserved isoforms in vertebrates, the invertebrate homologs are significantly less differentiated.