title: Glutamine's protection against cellular injury is dependent on heat shock factor-1.
abstract: Glutamine (GLN) has been shown to protect cells, tissues, and whole organisms from stress and injury. Enhanced expression of heat shock protein (HSP) has been hypothesized to be responsible for this protection. To date, there are no clear mechanistic data confirming this relationship. This study tested the hypothesis that GLN-mediated activation of the HSP pathway via heat shock factor-1 (HSF-1) is responsible for cellular protection. Wild-type HSF-1 (HSF-1(+/+)) and knockout (HSF-1(-/-)) mouse fibroblasts were used in all experiments. Cells were treated with GLN concentrations ranging from 0 to 16 mM and exposed to heat stress injury in a concurrent treatment model. Cell viability was assayed with phenazine methosulfate plus tetrazolium salt, HSP-70, HSP-25, and nuclear HSF-1 expression via Western blot analysis, and HSF-1/heat shock element (HSE) binding via EMSA. GLN significantly attenuated heat-stress induced cell death in HSF-1(+/+) cells in a dose-dependent manner; however, the survival benefit of GLN was lost in HSF-1(-/-) cells. GLN led to a dose-dependent increase in HSP-70 and HSP-25 expression after heat stress. No inducible HSP expression was observed in HSF-1(-/-) cells. GLN increased unphosphorylated HSF-1 in the nucleus before heat stress. This was accompanied by a GLN-mediated increase in HSF-1/HSE binding and nuclear content of phosphorylated HSF-1 after heat stress. This is the first demonstration that GLN-mediated cellular protection after heat-stress injury is related to HSF-1 expression and cellular capacity to activate an HSP response. Furthermore, the mechanism of GLN-mediated protection against injury appears to involve an increase in nuclear HSF-1 content before stress and increased HSF-1 promoter binding and phosphorylation.
authors.affiliation: Univ. of Colorado Health Sciences Center, Dept. of Anesthesiology, 4200 E. 9th Ave., Denver, CO 80262, USA.
authors.forename: Angela L
authors.initials: AL
authors.lastname: Morrison
authors.forename: Martin
authors.initials: M
authors.lastname: Dinges
authors.forename: Kristen D
authors.initials: KD
authors.lastname: Singleton
authors.forename: Kelli
authors.initials: K
authors.lastname: Odoms
authors.forename: Hector R
authors.initials: HR
authors.lastname: Wong
authors.forename: Paul E
authors.initials: PE
authors.lastname: Wischmeyer
mesh_terms: D000818:Animals; D015153:Blotting, Western; D002478:Cells, Cultured; D004268:DNA-Binding Proteins / Q000187:drug effects* / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D024202:Electrophoretic Mobility Shift Assay; D005347:Fibroblasts / Q000187:drug effects* / Q000473:pathology; D015870:Gene Expression; D005786:Gene Expression Regulation; D005973:Glutamine / Q000378:metabolism / Q000494:pharmacology*; D000076249:Heat Shock Transcription Factors; D018882:Heat Stress Disorders / Q000378:metabolism; D018869:Heat-Shock Response / Q000502:physiology*; D051379:Mice; D018345:Mice, Knockout; D014157:Transcription Factors / Q000187:drug effects* / Q000378:metabolism
chemical_list: D004268:DNA-Binding Proteins; D000076249:Heat Shock Transcription Factors; C490017:Hsf1 protein, mouse; D014157:Transcription Factors; D005973:Glutamine