title: Hypoxia stimulates migration of breast cancer cells via the PERK/ATF4/LAMP3-arm of the unfolded protein response.
abstract: BACKGROUND
  The hypoxia-inducible factor (HIF)-1 pathway can stimulate tumor cell migration and metastasis. Furthermore, hypoxic tumors are associated with a poor prognosis. Besides the HIF-1 pathway, the unfolded protein response (UPR) is also induced by hypoxic conditions. The PKR-like ER kinase (PERK)/activating transcription factor 4 (ATF4)-arm of the UPR induces expression of lysosomal-associated membrane protein 3 (LAMP3), a factor that has been linked to metastasis and poor prognosis in solid tumors. In this study the role of UPR-induced LAMP3 in hypoxia-mediated migration of breast cancer cells was examined.
  
  
  METHODS
  A number of in vitro metastasis models were used to study the migration and invasion of MDA-MB-231 breast cancer cells under hypoxic conditions. PERK, ATF4 and their downstream factor LAMP3 were knocked down to examine their role in cell migration. In addition, multicellular tumor spheroids were used to study the involvement of the tumor microenvironment in invasion.
  
  
  RESULTS
  Using transwell assays, migration of different breast cancer cell lines was assessed. A direct correlation was found between cell migration and baseline LAMP3 expression. Furthermore, moderate hypoxia (1% O2) was found to be optimal in stimulating migration of MDA-MB-231 cells. siRNA mediated knockdown of PERK, ATF4 and LAMP3 reduced migration of cells under these conditions. Using gap closure assays, similar results were found. In a three-dimensional invasion assay into collagen, LAMP3 knockdown cells showed a diminished capacity to invade compared to control cells when collectively grown in multicellular spheroids.
  
  
  CONCLUSIONS
  Thus, the PERK/ATF4/LAMP3-arm of the UPR is an additional pathway mediating hypoxia-induced breast cancer cell migration.
authors.forename: Anika
authors.initials: A
authors.lastname: Nagelkerke
authors.forename: Johan
authors.initials: J
authors.lastname: Bussink
authors.forename: Hilda
authors.initials: H
authors.lastname: Mujcic
authors.forename: Bradly G
authors.initials: BG
authors.lastname: Wouters
authors.forename: Steffi
authors.initials: S
authors.lastname: Lehmann
authors.forename: Fred C G J
authors.initials: FC
authors.lastname: Sweep
authors.forename: Paul N
authors.initials: PN
authors.lastname: Span
mesh_terms: D051701:Activating Transcription Factor 4 / Q000096:biosynthesis*; D001943:Breast Neoplasms / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D015687:Cell Hypoxia / Q000235:genetics; D045744:Cell Line, Tumor; D002465:Cell Movement / Q000235:genetics*; D005260:Female; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D051907:Lysosomal Membrane Proteins / Q000096:biosynthesis*; D009363:Neoplasm Proteins / Q000096:biosynthesis*; D034741:RNA, Small Interfering; D015398:Signal Transduction / Q000235:genetics; D056811:Unfolded Protein Response / Q000235:genetics; D019892:eIF-2 Kinase / Q000096:biosynthesis*; D059666:Lysosomal-Associated Membrane Protein 3
chemical_list: D051701:Activating Transcription Factor 4; D051907:Lysosomal Membrane Proteins; D009363:Neoplasm Proteins; D034741:RNA, Small Interfering; D019892:eIF-2 Kinase; C497109:ATF4 protein, human; C114233:LAMP3 protein, human; D059666:Lysosomal-Associated Membrane Protein 3; C460640:EIF2AK3 protein, human