Abstract
L-type amino acid transporter 1 (LAT1)/SLC7A5 is the first identified CD98 light chain disulfide linked to the CD98 heavy chain (CD98hc/SLC3A2). LAT1 transports large neutral amino acids, including leucine, which activates mTOR, and is highly expressed in human cancers. We investigated the oncogenicity of human LAT1 introduced to NIH/3T3 cells by retrovirus infection. NIH/3T3 cell lines stably expressing human native (164C) or mutant (164S) LAT1 (naLAT1/3T3 or muLAT1/3T3, respectively) were established. We confirmed that endogenous mouse CD98hc forms a disulfide bond with exogenous human LAT1 in naLAT1/3T3, but not in muLAT1/3T3. Endogenous mouse CD98hc mRNA increased in both naNIH/3T3 and muLAT1/3T3, and a similar amount of exogenous human LAT1 protein was detected in both cell lines. Furthermore, naLAT1/3T3 and muLAT1/3T3 cell lines were evaluated for cell growth-related phenotypes (phosphorylation of ERK, cell-cycle progression) and cell malignancy-related phenotypes (anchorage-independent cell growth, tumor formation in nude mice). naLAT1/3T3 had stronger growth- and malignancy- related phenotypes than NIH/3T3 and muLAT1/3T3, suggesting the oncogenicity of native LAT1 through its interaction with CD98hc. Anti-LAT1 monoclonal antibodies significantly inhibited in vitro cell proliferation and in vivo tumor growth of naLAT1/3T3 cells in nude mice, demonstrating LAT1 to be a promising anti-cancer target.
Highlights
CD98 was originally reported as a cell-surface marker associated with lymphocyte activation [1], and subsequently identified as a unique molecule expressed by numerous cancer cells [2,3,4]
We established NIH/3T3 cell lines infected with a vacant pMYs-IRES-Puro retrovirus, or the same retrovirus containing cDNA encoding human (h) native (164C) L-type amino acid transporter 1 (LAT1) or mutant (164 S) LAT1, whose 164th cysteine is genetically converted to serine
Regarding endogenous mouse LAT1 and CD98 heavy chain (CD98hc), the CD98hc mRNA level increased in both naLAT1/3T3 and muLAT1/3T3 compared with coNIH/3T3 cells, mLAT1 mRNA levels were almost equivalent among the three NIH/3T3 cell lines
Summary
CD98 was originally reported as a cell-surface marker associated with lymphocyte activation [1], and subsequently identified as a unique molecule expressed by numerous cancer cells [2,3,4]. Variant form of CD44 (CD44v) [12,13,14,15,16,17] and HER1 [18] associate with xCT in epithelial cancers or gliomas and stabilize xCT, respectively, resulting in the survival of CSCs in the oxidative stress by anti-cancer drugs. In this context, we demonstrated the antitumor effects of fully-human mAb [19] recognizing CD44v bound to xCT expressed by CSCs
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