The 3′UTR of the α6 integrin message regulates localization of α6β4 integrin heterodimers
Introduction
Cell adhesion and migration are critical processes for normal cell functioning, and these processes are aided by cell-matrix adhesion protein complexes: hemidesmosomes (HDs) and focal adhesions (FAs). The core of these complexes contains integrin heterodimers which interact with both the extracellular matrix and other cytoplasmic elements [1]. Each integrin heterodimer is a transmembrane, non-covalently bound complex that contains one α subunit and one β subunit. There are 18 α subunits and 8 β subunits which can form a variety of heterodimers, for example α2β1, α3β1, αvβ1, and α2β5. However, only 24 different heterodimers have been identified [2]. The α6β4 integrin heterodimer is unusual, not only because of the large size of the β4 integrin cytoplasmic tail, but also because it is only found in HDs and HD-related matrix adhesions, rather than in FAs [3]. HDs adhere epithelial cells to the extracellular matrix in several tissues including the skin, cornea, bladder, and some glands. In contrast, HD-related adhesions are found in certain alveolar cells in the lung and gut epithelial cells where they are presumed to mediate adhesion to the underlying connective tissue [[4], [5], [6]].
The α6 integrin subunit has two possible β subunit partners: β1 and β4 integrin. In cells which express both β1 and β4 integrin, α6 integrin preferentially pairs with β4, but how this is controlled is unclear [7,8]. Moreover, the targeting mechanism of α6β4 integrin to HD and HD-related matrix adhesion sites over other substrate attachment sites, including FAs, is also not understood. In this study, we have identified the 3′ untranslated region (UTR) of the α6 integrin messenger RNA (mRNA) as a key regulator of the localization of α6β4 integrin heterodimers as well as the formation of α6β4 versus α6β1 integrin heterodimers. In this regard, the 3′UTRs of mRNAs are known to regulate the shuttling of mRNAs to their site of translation by a sequence known as a zipcode. Although zipcode sequences can be located within coding regions of mRNA, generally, they are located within the 3′UTR [9]. For example, the zipcode in the β-actin message directs it to the site of FAs where it is translated [10]. Likewise, zipcodes in vinculin, actinin-4, and α3 integrin mRNAs direct their translation to the FA [11,12]. In this study we identified a region of the 3′UTR of α6 integrin mRNA which has zipcode characteristics and demonstrate the importance of the α6 integrin 3′UTR in regulating assembly and localization of α6β4 integrin heterodimers.
Section snippets
Cell culture conditions
A549s were cultured in HyClone MEM/EBSS media (HyClone, Logan, UT, USA) supplemented with 10% fetal bovine serum (Sigma-Aldrich, St. Louis, MO, USA), 2% l-glutamine, and 1% penicillin/streptomycin mixture. Immortalized human epithelial keratinocytes (iHEKs) were cultured in defined K-SFM keratinocyte media (DKSFM) (Thermo Fisher, Waltham, MA) supplemented with keratinocyte supplement and 1% penicillin/streptomycin mixture. Cells were maintained at 37 °C in 8% humidified CO2. iHEKs, exhibiting
Results and discussion
A zipcode within the α6 integrin 3′UTR is required for α6β4 heterodimer localization in A549 cells.
Zipcodes are sequences within 3′UTRs of mRNAs that direct their localization and subsequent translation. For example, the zipcode of α3 integrin was demonstrated to be necessary for α3 integrin protein translation and incorporation into FAs [12]. We therefore posited that the α6 integrin 3′UTR might be an important determinant of targeting α6β4 integrin heterodimers to HDs and HD-related
Conflicts of interest
No conflict of interests is noted by any of the listed authors.
Acknowledgements
We would like to thank Kevin Hamill of the University of Liverpool for helpful discussion. This work was begun while the senior author was a faculty member at the Feinberg School of Medicine, Northwestern University, Chicago, IL.
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