The vital importance of the Leloir pathway of
galactose metabolism has been repeatedly demonstrated by various uni-/multicellular model organisms, as well human patients who have inherited deficiencies of the key GAL
enzymes. Yet, other than the obvious links to the glycolytic pathway and
glycan biosynthetic pathways, little is known about how this metabolic pathway interacts with the rest of the metabolic and signaling networks. In this study, we compared the growth and the expression levels of the key components of the PI3K/Akt growth signaling pathway in primary fibroblasts derived from normal and galactose-1
phosphate uridylyltransferase (GalT)-deficient mice, the latter exhibited a
subfertility phenotype in adult females and growth restriction in both sexes. The growth potential and the
protein levels of the pAkt(Thr308), pAkt(Ser473), pan-Akt, pPdk1, and Hsp90
proteins were significantly reduced by 62.5%, 60.3%, 66%, 66%, and 50%, respectively in the GalT-deficient cells. Reduced expression of phosphorylated Akt
proteins in the mutant cells led to diminished phosphorylation of Gsk-3β (-74%).
Protein expression of BiP and pPten were 276% and 176% higher respectively in cells with
GalT-deficiency. Of the 24 genes interrogated using QIAGEN RT(2) Profiler PCR Custom Arrays, the
mRNA abundance of Akt1, Pdpk1, Hsp90aa1 and Pi3kca genes were significantly reduced at least 2.03-, 1.37-, 2.45-, and 1.78-fold respectively in mutant fibroblasts. Both serum-fasted normal and GalT-deficient cells responded to Igf-1-induced activation of Akt phosphorylation at +15 min, but the mutant cells have lower phosphorylation levels. The steady-state
protein abundance of
Igf-1 receptor was also significantly reduced in mutant cells. Our results thus demonstrated that
GalT deficiency can effect down-regulation of the PI3K/Akt growth signaling pathway in mouse fibroblasts through distinct mechanisms targeting both gene and
protein expression levels.