TY  - JOUR
A1  - Afonso-Grunz, Fabian
A1  - Hoffmeier, Klaus
A1  - Müller, Sören
A1  - Westermann, Alexander J.
A1  - Rotter, Björn
A1  - Vogel, Jörg
A1  - Winter, Peter
A1  - Kahl, Günter
T1  - Dual 3'Seq using deepSuperSAGE uncovers transcriptomes of interacting Salmonella enterica Typhimurium and human host cells
JF  - BMC Genomics
N2  - Background: 
The interaction of eukaryotic host and prokaryotic pathogen cells is linked to specific changes in the cellular proteome, and consequently to infection-related gene expression patterns of the involved cells. To simultaneously assess the transcriptomes of both organisms during their interaction we developed dual 3'Seq, a tag-based sequencing protocol that allows for exact quantification of differentially expressed transcripts in interacting pro-and eukaryotic cells without prior fixation or physical disruption of the interaction. 

Results: 
Human epithelial cells were infected with Salmonella enterica Typhimurium as a model system for invasion of the intestinal epithelium, and the transcriptional response of the infected host cells together with the differential expression of invading and intracellular pathogen cells was determined by dual 3'Seq coupled with the next-generation sequencing-based transcriptome profiling technique deepSuperSAGE (deep Serial Analysis of Gene Expression). Annotation to reference transcriptomes comprising the operon structure of the employed S. enterica Typhimurium strain allowed for in silico separation of the interacting cells including quantification of polycistronic RNAs. Eighty-nine percent of the known loci are found to be transcribed in prokaryotic cells prior or subsequent to infection of the host, while 75% of all protein-coding loci are represented in the polyadenylated transcriptomes of human host cells.

Conclusions: 
Dual 3'Seq was alternatively coupled to MACE (Massive Analysis of cDNA ends) to assess the advantages and drawbacks of a library preparation procedure that allows for sequencing of longer fragments. Additionally, the identified expression patterns of both organisms were validated by qRT-PCR using three independent biological replicates, which confirmed that RELB along with NFKB1 and NFKB2 are involved in the initial immune response of epithelial cells after infection with S. enterica Typhimurium.
KW  - complete genome sequence
KW  - secretion systems
KW  - RNA-Seq
KW  - deepSuperSAGE
KW  - transcriptome
KW  - gene expression
KW  - serovar Typhimurium
KW  - human macrophages
KW  - epithelial cells
KW  - infection
KW  - SuperSAGE
KW  - receptors
KW  - Dual 3'seq
KW  - MACE
KW  - tag based
KW  - simultaneous
KW  - genome wide
KW  - gene expression profiling
KW  - host pathogen interaction
KW  - Salmonella enterica Typhimurium strain SL1344
Y1  - 2015
U6  - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-143230
VL  - 16
IS  - 323
ER  -