Identity crisis in the PMP-22/EMP/MP20/Claudin superfamily (Pfam00822)

Abstract

We would like to call your attention to a publication by promotes an aggressive phenotype in human breast cancer cells") 1 in Tissue Barriers. The authors show that high claudin-20 expression correlates with the aggressiveness of breast cancer and poor survival rates for affected patients. They discuss a connection between the sensitivity of cancer cells to Gefitinib and claudin-20 content and suggest that claudin-20 might serve as a biomarker for Gefitinib resistance. However, this conclusion is based on a misconception on the part of the authors. They claim that claudin-20 is the same protein as epithelial membrane protein 1 (EMP-1), and to substantiate this claim, they misleadingly refer to a prior publication demonstrating that EMP-1 is a biomarker for Gefitinib-resistance in an adenocarcinoma model. However, Claudin-20 and EMP-1 are 2 distinct proteins. EMP-1 (NCBI reference sequence accession number NP_001414.1) has been described as a tight junction protein in the blood-brain barrier 3 as well as in the human airway epithelium, where it regulates tight junction formation. 4 To our knowledge, the paper of Martin et al. 1 is the first publication that presents functional data regarding human claudin-20 (NP_001001346.1). Their paper will be immediately found (and likely cited) by other scientists working with either of the 2 proteins. Therefore, it is crucial to avoid misrepresentations through a careful review of the existing literature.According to Price et al. 5 (also cited by Martin and colleagues 1 ) both claudin-20 and EMP-1 belong to the Pfam00822 superfamily, which includes claudins, peripheral myelin protein 22 (PMP-22), lens intrinsic membrane protein (LIM-2/MP20), EMP-1, ¡2, ¡3 and voltage dependent calcium channel gamma subunits (CACNGs). The study places claudin-20 within the claudin family (where it is most closely related to the classic claudins 2 and 14), while EMP-1 is placed in the LIM2/PMP22/EMP family.5 This classification was confirmed in an independent investigation, which came to the same results and confirmed the separate identities of claudin-20 and EMP-1. 6 Nevertheless, the phylogeny of the Pfam00822 family has not yet been completely resolved. TMEM114 and TMEM235 were first assigned to the claudin family using PMP-22 as an outgroup.7 Later, these TMEMs were renamed claudin-26 and ¡27, respectively. 8 Subsequent work that also included CACNGs, EMPs and LIM-2 and used clarin-1 as an outgroup revealed that TMEM114 and TMEM235 were more closely related to the CACNG family than with the claudin family. 6 Thus, Pfam00822 superfamily nomenclature has not yet been conclusively determined.The members of this superfamily are tetraspanning membrane proteins and share a WX 9-44 GLWXXC(X 9-26 C) signature motif in the first extracellular loop (whereas the second cysteine is not conserved in the LIM2/PMP22/EMP family). To illustrate the differences between EMP-1 and claudin-20, we modeled their structures using I-TASSER. 9 The recently published crystal structure of claudin-15 10 was applied a...

A face-to-face comparison of claudin-5 transduced human brain endothelial (hCMEC/D3) cells with porcine brain endothelial cells as blood–brain barrier models for drug transport studies

“…First, meninges and large blood vessels were removed from the cerebral cortex under sterile conditions. After roughly separating white from gray matter, the gray matter was minced and sequentially treated with digestion enzymes, density gradient centrifugation, filtration and erythrocyte lysing as described earlier [ 19 ]. Purified pBCECs were seeded in the apical compartment of collagen IV-coated Transwell ® chambers (12 well format, 0.4 µm pore size, 1.12 cm 2 growth area, polyester membrane, transparent, Corning Costar, #3460) at 37 °C and 5% CO 2 in a density of 3.6 × 10 5 cells/cm 2 in medium 199 (Gibco/Life Technologies, Carlsbad, CA, USA) supplemented with 10% newborn calf serum (Biochrom), 0.7 mM l -glutamine (Gibco/Life Technologies), 1% penicillin/streptomycin, and 1% gentamicin (Gibco/Life Technologies).…”

“…Differences are mainly localized in transmembrane domain 3 and 4. However, molecular modelling studies show that the differences do not influence their helical structure of the domains and, hence, the claudin-5 function in the junction [19,20]. Important for the barrier function of claudin-5 is its paracellular tightening activity, which is caused by its extracellular domain consisting of the two extracellular loops (ECLs) [21].…”

“…First, meninges and large blood vessels were removed from the cerebral cortex under sterile conditions. After roughly separating white from gray matter, the gray matter was minced and sequentially treated with digestion enzymes, density gradient centrifugation, filtration and erythrocyte lysing as described earlier [19]. Purified pBCECs were Fig.…”