Affymetrix GeneChip Yeast Genome 2.0 Array [Yeast_2]

Glucose depletion inhibits translation initiation via eIF4A loss and subsequent 48S preinitiation complex accumulation, while the pentose phosphate pathway is coordinately up-regulated

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This is a polysome/monosome profiling study on yeast undergoing glucose depletion. Polysomes and monosomes are extracted and the transcripts bound to each fraction are measured by microarray. The study reveals that glucose depletion inhibits translation initiation in a mechanism involving eIF4A loss and 48S pre-initiation complex accumulation, while the pentose phosphate pathway is co-ordinately up-regulated

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Glucose depletion inhibits translation initiation in yeast - polysome and monosome profiling

This is a polysome/monosome profiling study on yeast undergoing glucose depletion. Polysomes and monosomes are extracted and the transcripts bound to each fraction are measured by microarray. The study reveals that glucose depletion inhibits translation initiation in a mechanism involving eIF4A loss and 48S pre-initiation complex accumulation, while the pentose phosphate pathway is co-ordinately up-regulated


Affymetrix:Protocol:Hybridization-Unknown

P-MTAB-18358

Polysome microarrays were performed as described previously (Smirnova et al, Molecular and Cellular Biology, November 2005, p. 9340-9349, Vol. 25, No. 21 0270-7306/05/$08.00+0). Briefly, polysomes and monosomes are separated and the transcripts bound to each fraction are measured by microarray. Cycloheximide treated chilled cultures were snap frozen in liquid nitrogen and cells were lysed by grinding under liquid nitrogen.  Lysates were cleared by centrifugation at 5000 rpm at 4°C for 1 hour.  One hundred A260 units were loaded onto 35ml 15 to 50% sucrose gradients. The gradients were sedimented via centrifugation at 16,900 rpm in an SW28 rotor (Beckman) for 13 h.  The gradients were collected using an ISCO gradient collection and fractionation system adapted for the larger 1 x 3.5 inch centrifuge tubes and fractionated into 15 x 2.2ml aliquots. These aliquots were collected directly into 2 volumes of Trizol (Invitrogen) and stored at -80°C.  Fractions 4-8 (monosomal peaks) and 11-15 (polysomal peaks) were pooled for the subsequent RNA extractions. 

P-MTAB-18359

P-MTAB-18360

P-MTAB-18361

Background correction,quantile normalization, and gene expression analysis were performed using RMA (Bolstad, B.M., Irizarry R. A., Astrand, M., and Speed, T.P. (2003)A Comparison of Normalization Methods for High Density Oligonucleotide Array Data Based on Bias and Variance. Bioinformatics 19(2):185-193).

Glucose depletion inhibits translation initiation in yeast - polysome and monosome profiling

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