title: The PI3K/Akt1 pathway enhances steady-state levels of FANCL.
abstract: Fanconi anemia hematopoietic stem cells display poor self-renewal capacity when subjected to a variety of cellular stress. This phenotype raises the question of whether the Fanconi anemia proteins are stabilized or recruited as part of a stress response and protect against stem cell loss. Here we provide evidence that FANCL, the E3 ubiquitin ligase of the Fanconi anemia pathway, is constitutively targeted for degradation by the proteasome. We confirm biochemically that FANCL is polyubiquitinated with Lys-48-linked chains. Evaluation of a series of N-terminal-deletion mutants showed that FANCL's E2-like fold may direct ubiquitination. In addition, our studies showed that FANCL is stabilized in a complex with axin1 when glycogen synthase kinase-3β is overexpressed. This result leads us to investigate the potential regulation of FANCL by upstream signaling pathways known to regulate glycogen synthase kinase-3β. We report that constitutively active, myristoylated-Akt increases FANCL protein level by reducing polyubiquitination of FANCL. Two-dimensional PAGE analysis shows that acidic forms of FANCL, some of which are phospho-FANCL, are not subject to polyubiquitination. These results indicate that a signal transduction pathway involved in self-renewal and survival of hematopoietic stem cells also functions to stabilize FANCL and suggests that FANCL participates directly in support of stem cell function.
authors.affiliation: Knight Cancer Institute, Oregon Health and Science University, Portland, OR 97239, USA. daok@ohsu.edu
authors.forename: Kim-Hien T
authors.initials: KH
authors.lastname: Dao
authors.forename: Michael D
authors.initials: MD
authors.lastname: Rotelli
authors.forename: Brieanna R
authors.initials: BR
authors.lastname: Brown
authors.forename: Jane E
authors.initials: JE
authors.lastname: Yates
authors.forename: Juha
authors.initials: J
authors.lastname: Rantala
authors.forename: Cristina
authors.initials: C
authors.lastname: Tognon
authors.forename: Jeffrey W
authors.initials: JW
authors.lastname: Tyner
authors.forename: Brian J
authors.initials: BJ
authors.lastname: Druker
authors.forename: Grover C
authors.initials: GC
authors.lastname: Bagby
mesh_terms: D060466:Axin Protein / Q000235:genetics / Q000378:metabolism; D002460:Cell Line; D004789:Enzyme Activation; D005199:Fanconi Anemia / Q000378:metabolism; D052240:Fanconi Anemia Complementation Group L Protein / Q000235:genetics / Q000378:metabolism*; D015870:Gene Expression; D038362:Glycogen Synthase Kinase 3 / Q000096:biosynthesis / Q000235:genetics; D000071679:Glycogen Synthase Kinase 3 beta; D057809:HEK293 Cells; D006367:HeLa Cells; D006412:Hematopoietic Stem Cells / Q000378:metabolism*; D006801:Humans; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism*; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism; D017510:Protein Folding; D051057:Proto-Oncogene Proteins c-akt / Q000235:genetics / Q000378:metabolism*; D034622:RNA Interference; D034741:RNA, Small Interfering; D015398:Signal Transduction; D054875:Ubiquitination
chemical_list: C555451:AXIN1 protein, human; D060466:Axin Protein; D034741:RNA, Small Interfering; C499908:FANCL protein, human; D052240:Fanconi Anemia Complementation Group L Protein; D019869:Phosphatidylinositol 3-Kinases; C494918:AKT1 protein, human; C000605506:GSK3B protein, human; D000071679:Glycogen Synthase Kinase 3 beta; D051057:Proto-Oncogene Proteins c-akt; D038362:Glycogen Synthase Kinase 3; D046988:Proteasome Endopeptidase Complex