PMID- 30029480
OWN - NLM
STAT- MEDLINE
DCOM- 20181029
LR  - 20190220
IS  - 1422-0067 (Electronic)
IS  - 1422-0067 (Linking)
VI  - 19
IP  - 7
DP  - 2018 Jul 19
TI  - Targeted Suppression and Knockout of ASCT2 or LAT1 in Epithelial and Mesenchymal 
      Human Liver Cancer Cells Fail to Inhibit Growth.
LID - 10.3390/ijms19072093 [doi]
LID - 2093
AB  - Amino acid transporters alanine-serine-cysteine transporter 2 (ASCT2) and L-Type 
      Amino Acid Transporter 1 (LAT1) are coordinately enhanced in human cancers where 
      among other roles, they are thought to drive mechanistic target-of-rapamycin 
      (mTOR) growth signaling. To assess ASCT2 and LAT1 as therapeutic targets, nine 
      unique short hairpin RNA (shRNA) vectors were used to stably suppress transporter 
      expression in human epithelial (Hep3B) and mesenchymal (SK-Hep1) hepatocellular 
      carcinoma (HCC) cell lines. In addition, six unique CRISPR-Cas9 vectors were used 
      to edit the ASCT2 (SLC1A5) and LAT1 (SLC7A5) genes in epithelial (HUH7) and 
      mesenchymal (SK-Hep1) HCC cells. Both approaches successfully diminished 
      glutamine (ASCT2) and leucine (LAT1) initial-rate transport proportional to 
      transporter protein suppression. In spite of profoundly reduced glutamine or 
      leucine transport (up to 90%), transporter suppression or knockout failed to 
      substantially affect cellular proliferation or basal and amino acid-stimulated 
      mTORC1 growth signaling in either HCC cell type. Only LAT1 knockout in HUH7 
      slightly reduced growth rate. However, intracellular accumulation of radiolabeled 
      glutamine and leucine over longer time periods largely recovered to control 
      levels in ASCT2 and LAT1 knockout cells, respectively, which partially explains 
      the lack of an impaired growth phenotype. These data collectively establish that 
      in an in vitro context, human epithelial and mesenchymal HCC cell lines adapt to 
      ASCT2 or LAT1 knockout. These results comport with an emerging model of amino 
      acid exchangers like ASCT2 and LAT1 as "harmonizers", not drivers, of amino acid 
      accumulation and signaling, despite their long-established dominant role in 
      initial-rate amino acid transport.
FAU - Bothwell, Paige J
AU  - Bothwell PJ
AD  - Department of Biological Sciences/Center for Biochemical and Biophysical Studies, 
      Northern Illinois University, DeKalb, IL 60115, USA. pbothwell1@niu.edu.
FAU - Kron, Clare D
AU  - Kron CD
AD  - Department of Biological Sciences/Center for Biochemical and Biophysical Studies, 
      Northern Illinois University, DeKalb, IL 60115, USA. clarekron@frontier.com.
FAU - Wittke, Evan F
AU  - Wittke EF
AUID- ORCID: 0000-0001-8482-0636
AD  - Department of Biological Sciences/Center for Biochemical and Biophysical Studies, 
      Northern Illinois University, DeKalb, IL 60115, USA. ewittke@luc.edu.
FAU - Czerniak, Bradley N
AU  - Czerniak BN
AD  - Department of Biological Sciences/Center for Biochemical and Biophysical Studies, 
      Northern Illinois University, DeKalb, IL 60115, USA. bczerniak2@hotmail.com.
FAU - Bode, Barrie P
AU  - Bode BP
AD  - Department of Biological Sciences/Center for Biochemical and Biophysical Studies, 
      Northern Illinois University, DeKalb, IL 60115, USA. bodebp@niu.edu.
LA  - eng
GR  - T35 HL120835/HL/NHLBI NIH HHS/United States
PT  - Journal Article
DEP - 20180719
PL  - Switzerland
TA  - Int J Mol Sci
JT  - International journal of molecular sciences
JID - 101092791
RN  - 0 (Amino Acid Transport System ASC)
RN  - 0 (Amino Acids)
RN  - 0 (Large Neutral Amino Acid-Transporter 1)
RN  - 0 (Minor Histocompatibility Antigens)
RN  - 0 (RNA, Antisense)
RN  - 0 (RNA, Small Interfering)
RN  - 0 (SLC1A5 protein, human)
RN  - 320T6RNW1F (Mifepristone)
RN  - 9NEZ333N27 (Sodium)
RN  - EC 2.7.11.1 (Mechanistic Target of Rapamycin Complex 1)
SB  - IM
MH  - Amino Acid Transport System ASC/*metabolism
MH  - Amino Acids/metabolism
MH  - Biological Transport/drug effects
MH  - CRISPR-Cas Systems/genetics
MH  - Cell Death/drug effects
MH  - Cell Proliferation/drug effects
MH  - Epithelium/*pathology
MH  - *Gene Knockout Techniques
MH  - Humans
MH  - Large Neutral Amino Acid-Transporter 1/*metabolism
MH  - Liver Neoplasms/*metabolism/*pathology
MH  - Mechanistic Target of Rapamycin Complex 1/metabolism
MH  - Mesoderm/*pathology
MH  - Mifepristone/pharmacology
MH  - Minor Histocompatibility Antigens/*metabolism
MH  - RNA, Antisense/metabolism
MH  - RNA, Small Interfering/metabolism
MH  - Signal Transduction/drug effects
MH  - Sodium/metabolism
PMC - PMC6073291
OTO - NOTNLM
OT  - ASCT2
OT  - CRISPR-Cas9
OT  - LAT1
OT  - SLC1A5
OT  - SLC7A5
OT  - amino acid transport
OT  - hepatocellular carcinoma
OT  - mTOR
OT  - shRNA
COIS- The authors declare no conflicts of interest.
EDAT- 2018/07/22 06:00
MHDA- 2018/10/30 06:00
PMCR- 2018/07/01
CRDT- 2018/07/22 06:00
PHST- 2018/06/25 00:00 [received]
PHST- 2018/07/05 00:00 [accepted]
PHST- 2018/07/22 06:00 [entrez]
PHST- 2018/07/22 06:00 [pubmed]
PHST- 2018/10/30 06:00 [medline]
PHST- 2018/07/01 00:00 [pmc-release]
AID - ijms19072093 [pii]
AID - ijms-19-02093 [pii]
AID - 10.3390/ijms19072093 [doi]
PST - epublish
SO  - Int J Mol Sci. 2018 Jul 19;19(7):2093. doi: 10.3390/ijms19072093.