Cingulin (CGN; from the Latin cingere “to form a belt around”) is a cytosolic
protein encoded by the CGNgene in humans[5][6][7] localized at
tight junctions (TJs) of vertebrate epithelial and endothelial cells.
Discovery
Cingulin was originally discovered at the MRC Laboratory of Molecular Biology (Cambridge, UK) by Dr. Sandra Citi, as a protein present in chicken intestinal epithelial cells, that co-purified with non-muscle
myosin II and was specifically localized at tight junctions (zonulae occludentes).[8]
Structure & interactions
Cingulin is a homodimer, each subunit containing a N-terminal globular "head" domain, a long α-helical coiled-coil "rod" domain and a small globular C-terminal "tail" region.[9] This organization is highly conserved throughout
vertebrates.[5] However, cingulin
homologs have not been detected in
invertebrates.
In vitro, cingulin can bind to and bundle actin filaments, and interact with myosin II and several TJ proteins including ZO-1, ZO-2, ZO-3,
paracingulin and
occludin.[10][11][12] Moreover, cingulin forms a complex with JAM-A, a tight junction membrane protein.[10] Most of cingulin protein interactions are through the globular head domain. Cingulin interacts with ZO-1 through an N-terminal ZO-1 interacting motif (ZIM) in its head region.[13][14] The rod domain is involved in dimerization and interaction with the RhoA activator, GEF-H1.[15][16][17]
Cingulin has also been found to interact with microtubules (MTs) through the N-terminal head region, and these interactions was regulated by phosphorylation by the adenosine monophosphate-activated protein kinase (AMPK).[18]
Function
The function of cingulin has been studied by
knockout (KO),
knockdown (KD) and over-expression approaches. Embryoid bodies derived from embryonic stem cells where one or both cingulin alleles were targeted by homologous recombination show apparently normal tight junctions, but changes in the expression of a large number of genes, including tight junction protein genes (
claudin-2,
claudin-6,
claudin-7 and
occludin) and transcription factors (including
GATA4).[13] Changes in the expression of claudin-2 and ZO-3 are also observed in cultured kidney cells (MDCK) depleted of cingulin by
shRNA.[16]
In 2012, the phenotype of cingulin-knockout mice was described, proving that functional TJ in vivo can be formed in the absence of cingulin.[19] Together with paracingulin, cingulin also was reported to regulate
claudin-2 expression through RhoA-dependent and independent mechanisms.[19][20]
The role of cingulin in development has been studied by
morpholino.[21] oligonucleotide-mediated depletion in chicken, indicating that cingulin is involved in neural crest development. In early mouse and frog embryos, maternal cingulin is localized in the cell cortex. Through early mouse development, cytocortical cingulin in present from oogenesis (cumulus-oocyte contact sites) until 16-cells morulae stage (apical microvillous zones) during early embryogenesis; then maternal cingulin is degraded by endocytic turn-over from the 32-cells stage. Regarding the
zygotic cingulin, it accumulates at the tight junctions from 16-cells stage, 10 hours after ZO-1 assembly. Furthermore, the synthesis of cingulin in early mouse embryos is tissue-specific and it occurs in blastocyst (up-regulated in
trophectoderm and down-regulated in inner-cells).[22][23] In Xenopuslaevis embryos, maternal cingulin is recruited to apical cell-cell junctions from 2-cells stage.[24][25]
Homologs
In 2004, a protein homologous to cingulin was discovered and named JACOP (also known as paracingulin, or cingulin-like 1 protein;
CGNL1).[17]
Human diseases
Although cingulin has been involved in regulation of RhoA signaling and gene expression in cultured cells and KO mice, nothing is known about the specific role of cingulin in human diseases.[15][16][19]
Cingulin expression has been studied in human carcinomas and shown to be expressed in adenocarcinomas and down-regulated in squamous carcinomas.[26][27] Furthermore, histone deacetylase inhibitors, such as sodium butyrate, strongly upregulate its expression in some cultured cells.[28] Cingulin, as other junctional proteins could be used as a marker of epithelial differentiation, and as a diagnostic marker to distinguish adenocarcinomas from squamous carcinomas.
^"Human PubMed Reference:". National Center for Biotechnology Information, U.S. National Library of Medicine.
^"Mouse PubMed Reference:". National Center for Biotechnology Information, U.S. National Library of Medicine.
^
abCiti S, D'Atri F, Parry DA (August 2000). "Human and Xenopus cingulin share a modular organization of the coiled-coil rod domain: predictions for intra- and intermolecular assembly". Journal of Structural Biology. 131 (2): 135–45.
doi:
10.1006/jsbi.2000.4284.
PMID11042084.
^
abGuillemot L, Citi S (2006). "Cingulin, a Cytoskeleton-Associated Protein of the Tight Junction". In Gonzalez-Mariscal L (ed.). Tight junctions. Georgetown, Texas: Landes Bioscience/Eurekah.com. pp. 54–63.
ISBN978-0-387-36673-9.
^Kos R, Reedy MV, Johnson RL, Erickson CA (April 2001). "The winged-helix transcription factor FoxD3 is important for establishing the neural crest lineage and repressing melanogenesis in avian embryos". Development. 128 (8): 1467–79.
doi:
10.1242/dev.128.8.1467.
PMID11262245.
^Javed Q, Fleming TP, Hay M, Citi S (March 1993). "Tight junction protein cingulin is expressed by maternal and embryonic genomes during early mouse development". Development. 117 (3): 1145–51.
doi:
10.1242/dev.117.3.1145.
PMID8325239.
^Fleming TP, Hay M, Javed Q, Citi S (March 1993). "Localisation of tight junction protein cingulin is temporally and spatially regulated during early mouse development". Development. 117 (3): 1135–44.
doi:
10.1242/dev.117.3.1135.
PMID8325238.
Gevaert K, Goethals M, Martens L, Van Damme J, Staes A, Thomas GR, Vandekerckhove J (May 2003). "Exploring proteomes and analyzing protein processing by mass spectrometric identification of sorted N-terminal peptides". Nature Biotechnology. 21 (5): 566–9.
doi:
10.1038/nbt810.
PMID12665801.
S2CID23783563.
Kim JE, Tannenbaum SR, White FM (2005). "Global phosphoproteome of HT-29 human colon adenocarcinoma cells". Journal of Proteome Research. 4 (4): 1339–46.
doi:
10.1021/pr050048h.
PMID16083285.