Better explicating the strengths and shortcomings of these models will help refine projections and improve transparency in the years ahead.
Image credit: Witsawat.S.
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Communicated by Allan Campbell, Stanford University, Stanford, CA (received for review October 29, 2001)
Article Figures & SI
Figures
- Figure 1
Time course effect of biotin deficiency on mRNA levels. HepG2 cells were grown in a biotin-free medium. Total RNA was isolated at different times and HCS, ACC-1, ACC-2, and PCCA mRNAs were amplified by reverse transcription PCR and quantitated by densitometry as described in Materials and Methods. (A) representative experiment showing the effect of time of biotin restriction on the mRNA levels for HCS, ACC-1, ACC-2, and PCCA. (B) Summary of the results obtained in three different experiments presented as mean ± SE. Lane numbers correspond to days in biotin-deficient medium.
- Figure 2
Effect of biotin and cGMP on mRNA levels. HepG2 cells cultivated 14 days in a biotin-free medium were stimulated with 1 μM biotin or 1 mM 8-Br-cGMP. HCS, ACC-1, ACC-2, PCCA, and β-actin mRNA were determined as described in Materials and Methods. (A) Representative experiment showing mRNA levels in biotin-replete cells (r), biotin-starved cells (s), biotin-starved cells stimulated with biotin (s + bio), and biotin-starved cells stimulated with 8-Br-cGMP (s + cGMP). (B) Summary of the results obtained from three different experiments. Results are presented as mean ± SE.
- Figure 3
Effect of biotin concentration on HCS mRNA levels in normal and MCD fibroblasts. Normal and MCD cells were grown in biotin free medium for 14 days. After this period, the cells were stimulated with 0.01 μM, 0.1 μM, and 1.0 μM biotin for 24 h. HCS mRNA levels were determined as described in Materials and Methods. Results are presented as percentage of mRNA values observed in cells grown in biotin-containing medium and normalized to β-actin mRNA levels. Data are from three different experiments and shown as mean ± SE.
Tables
- Table 1
Effect of inhibition of soluble guanylate cyclase, cGMP-dependent protein kinase, and RNA polymerase II in mRNA levels
mRNA Bio Def, % Bio Def + Bio, % ODQ + Bio, % PKGi + Bio, % Act D + Bio, % HCS 36 ± 5.0 103 ± 9.87 11 ± 0.68 52 ± 2.51 25 ± 6.0 ACC-1 34 ± 1.15 140 ± 2.21 36 ± 5.0 26 ± 9.53 14 ± 7.24 PCCA 38 ± 3.54 130 ± 6.61 33 ± 3.60 21 ± 11.1 34 ± 5.03 ACC-2 88 ± 8.96 95 ± 2.31 ND ND 19 ± 2.0 -
Biotin-starved HepG2 cells were stimulated with biotin in the presence of ODQ (ODQ + Bio), Rp-cGMPS (PKGi + Bio), or actinomycin-D. The results are shown as percentage of values obtained in cells grown in normal medium and compared to biotin-deficient cells (Bio Def) and biotin-deficient cells stimulated with biotin without inhibitors (Bio Def + Bio). ND, not determined. Results are from three different experiments shown as mean ± SE.
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- Table 2
Effect of different concentrations of cGMP on HCS mRNA levels in normal and MCD cells
Fibroblasts Bio Def, % Bio Def + cGMP, % Normal 34 ± 7.6 117 ± 1.5 MCD-MK 38 ± 12.6 92 ± 9.24 -
Biotin-starved MCD cells and normal fibroblasts were stimulated with 1.0 mM 8-Br-cGMP. Data are from three different experiments and are shown as mean ± SE.
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Holocarboxylase synthetase is an obligate participant in biotin-mediated regulation of its own expression and of biotin-dependent carboxylases mRNA levels in human cells
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