Naslov
Differences in endocyic pathway of CD44 molecule in comparison with other molecules

Autori
Karleuša, Ljerka ; Mahmutefendić, Hana ; Blagojević Zagorac, Gordana ; Ilić Tomaš, Maja ; Lučin, Pero

Vrsta, podvrsta i kategorija rada
Sažeci sa skupova, sažetak, znanstveni

Skup
2nd Meeting of Middle-European Societies for Immunology and Allergology

Mjesto i datum
Opatija, Hrvatska, 10.10.2013. - 13.10.2013

Vrsta sudjelovanja
Poster

Vrsta recenzije
Međunarodna recenzija

Ključne riječi
CD44; endocytosis; MHC class I

Sažetak
Introduction: CD44 molecule, a receptor for hyaluronic acid, is present at the cell surface in lipid-ordered and lipid-disordered plasma membrane (PM) environment. Although, it has been shown that CD44 associates with non-clathrin Arf6-associated endosomal carriers, similar to Major Histocompatibility Class I (MHC-I) proteins, its membrane turnover, internalization route and endosomal trafficking still remains poorly charachterized. Objectives: In this research we further investigated the internalization, recycling and degradation pathways of CD44 by comparing it with molecules with better elucidated endocytic pathways. Matherials and Methods: CD44 together with other molecules (fully conformed Dd, open forms of Ld, transferrin receptor and GM1 receptor) were followed by corresponding monoclonal antibodies or fluorescent labeled ligands on Balb 3T3 fibroblasts. Internalization rates of cell surface proteins were determined by flow cytometry using the internalization assay based on mAb labeling at the cell surface. The intracellular routes of the same proteins were determined by immunofluorescence and confocal analysis using various modifications of the pulse-chase internalization assay. In order to clarify the endocytic pathways of CD44 we also used inhibitor AlF4- and colocalization of internalized proteins with markers of endosomal compartments and pathways. Results: CD44 molecules localized in plasma membrane (PM) tubular invaginations. The internalization rate of CD44 was low and the cell surface half-life was long, indicating its high recycling rate that takes place via long PM tubular invaginations. Internalized CD44 appears to be contained in the vesicular formations that vary in size and placement when in cytoplasm. Very little of internalized CD44 colocalized with EEA1 and the colocalization with transferrin was almost non-existant meaning that almost no CD44 was detected in the juxanuclear recycling compartment. CTxB and CD44 molecules were found partially colocalized and partially comaprtmentalized in the cytoplasmatic regions close to the PM. There was very little to none colocalization of internalized CD44 and DND as well as CD44 and LBPA. This shows that CD44 rarely enters the late endosomes. The colocalization pattern form slight to very little has been observed also when comparing internalizations of CD44 and open forms of Ld as well as CD44 and fully conformed Dd molecules. Aluminum fluoride (AlF4-) increased the intercellular retention of CD44, same as to the MHC-I molecules, confirming our conclusion that peripheral recycling maintains the low internalization rate. Finally, the pH disrupting drug Concanamycin A did not change the internalization rate of CD44, whereas increased internalization of MHC-I molecules, suggesting that CD44 is retained in the pH neutral environment and internalized via neutral endosomal carriers. Conclusion: Long cell surface half-life of the CD44 molecules is probably maintained by high rate of peripheral recycling via long plasma membrane tubular invaginations. The endocytic route of CD44 is noticeably distinct from the one that is typical to the one of MHC-I molecules although some entouched subplasmalaemal and periphareal tubules could be observed. Internalized CD44 molecules rarely enter the juxanuclear recycling compartment. and seem not to enter late endosomes as well. They share slight endocytotic pathway with GM1 receptor, mostly in the cytoplasmatic regions close to the PM. According to that, CD44 follows endocytic pathway that is prevalently distinct from classical endocytic routes followed by transferrin receptor, GM1receptor and MHC-I molecules, although some minor tubular overlapping exists.

Izvorni jezik
Engleski

Znanstvena područja
Temeljne medicinske znanosti

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