Transcriptome profiling of genes involved in neural tube closure during human embryonic development using long serial analysis of gene expression (long-SAGE).
BACKGROUND
Neural tube defects (NTDs) are common human birth defects with a complex etiology. To develop a comprehensive knowledge of the genes expressed during normal neurulation, we established transcriptomes from human neural tube fragments during and after neurulation using long Serial Analysis of Gene Expression (long-SAGE).


METHODS
Rostral and caudal neural tubes were dissected from normal human embryos aged between 26 and 32 days of gestation. Tissues from the same region and Carnegie stage were pooled (n ≥ 4) and total RNA extracted to construct four long-SAGE libraries. Tags were mapped using the UniGene Homo sapiens 17 bp tag-to-gene best mapping set. Differentially expressed genes were identified by chi-square or Fisher's exact test, and validation was performed for a subset of those transcripts using in situ hybridization. In silico analyses were performed with BinGO and EXPANDER.


RESULTS
We observed most genes to be similarly regulated in rostral and caudal regions, but expression profiles differed during and after closure. In silico analysis found similar enrichments in both regions for biologic process terms, transcription factor binding and miRNA target motifs. Twelve genes potentially expressing alternate isoforms by region or developmental stage, and the microRNAs miR-339-5p, miR-141/200a, miR-23ab, and miR-129/129-5p are among several potential candidates identified here for future research.


CONCLUSIONS
Time appears to influence gene expression in the developing central nervous system more than location. These data provide a novel complement to traditional strategies of identifying genes associated with human NTDs and offer unique insight into the genes associated with normal human neurulation.
Center for Human Genetics, Department of Medicine, Duke University Medical Center, Durham, North Carolina 27710, USA.
Deidre R
DR
Krupp
Pu-Ting
PT
Xu
Sophie
S
Thomas
Andrew
A
Dellinger
Heather C
HC
Etchevers
Michel
M
Vekemans
John R
JR
Gilbert
Marcy C
MC
Speer
Allison E
AE
Ashley-Koch
Simon G
SG
Gregory
D000029:Abortion, Legal; D002874:Chromosome Mapping; D003198:Computer Simulation; D004622:Embryo, Mammalian; D005260:Female; D020869:Gene Expression Profiling; D018507:Gene Expression Regulation, Developmental; D015698:Genomic Library; D006801:Humans; D017403:In Situ Hybridization; D035683:MicroRNAs / Q000235:genetics*; D008957:Models, Genetic; D054259:Neural Tube / Q000166:cytology / Q000378:metabolism*; D009436:Neural Tube Defects / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D054261:Neurulation / Q000235:genetics*; D020411:Oligonucleotide Array Sequence Analysis; D011247:Pregnancy; D012333:RNA, Messenger / Q000235:genetics*; D013997:Time Factors; D014158:Transcription, Genetic
D035683:MicroRNAs; D012333:RNA, Messenger