Identification of Switching-Related Transcription Factors from a Comparison of Cord and Adult Human Blood Reticulocytes Transcriptomes.

Hemoglobin switching patterns during human ontogeny are highly correlated with transcription of the alpha and beta globin genes on chromosomes 16 and 11, but the molecular and genetic mechanisms responsible for the switching phenomenon are not yet defined. For a better understanding of post-natal globin gene switching and silencing, the expression profiles from purified reticulocyte mRNA (28 separate clinical samples; 14 cord, 14 adult) were studied using Affymetrix HG-U133 arrays (44,229 probe sets). To validate the transcriptome expression levels derived from the microarrays and to discriminate the differences between cord blood and adult blood reticulocytes, we performed quatitative real-time PCR. Among 21 tested control genes, the ratios of adult-to-cord blood (AB/CB) array hybridization intensities versus PCR copy numbers demonstrated a correlation coefficient of 0.978. Hence, the AB/CB array hybridization intensities provide a reliable description of erythroid transcriptome patterns associated with post-natal hemoglobin switching. Stringent screening criteria were then used to identify genes that may be involved in fetal-to-adult hemoglobin switching. All 44,229 probe sets were filtered to identify genes that demonstrated greater than a five fold difference in AB and CB mean signal intensities as well as consistent differences among the 14 samples from each group. In addition, the screening criteria required high-level signals in either AB or CB (>1000 FU), and t -test p -values between AB and CB