Skip to main content
Premium Trial:

Request an Annual Quote

Simultaneous RNA-seq of Host and Bacterial Invader Offer Glimpse into Disease Processes

NEW YORK (GenomeWeb News) – By turning to a simultaneous RNA-seq strategy, researchers at the University of Maryland School of Medicine profiled the transcriptomes of Chlamydia trachomatis and the host cells the bacteria infected, gaining a glimpse into how disease is established and causes harm.

As they reported in PLOS One yesterday, the researchers found that the host cells react quickly to C. trachomatis infection. However, host response to such infection contributes to scarring and other effects that are the hallmarks of chlamydia infection. By sifting through the transcriptomes, the Maryland team identified a possible positive feedback mechanism concentrated around the extracellular matrix- and tissue remodeling-related genes tenascin C, gremlin1, and TGF-β that may lead to scarring.

"We found that the response to chlamydial infection is rapid and dramatic, and observed many novel host cell transcriptional reactions to the infection," said Garry Myers, an assistant professor at the University of Maryland School of Medicine's Institute for Genome Sciences, in a statement. "In particular, we were able to identify abnormal early expression of host cell genes known to induce scarring and expression of numerous genes that encode the building blocks of fibrotic scars."

Scarring and tissue damage caused by chlamydial infection can lead to infertility in women if the fallopian tubes are affected and to trachoma or infectious blindness if the eyelids are affected.

To examine the interplay between C. trachomatis and its host cells, Myers and his colleagues infected an epithelial cell line with the bacteria, and extracted total RNA from them and from a set of mock-infected controls one hour and 24 hours after infection.

That RNA was then split into two portions, both of which underwent rRNA depletion, but one of the two groups was subject to an additional poly-A tail subtraction to increase the levels of bacterial mRNA. The samples were sequenced using the Illumina HiSeq2000 platform and mapped to the C. trachomatis reference genome using Bowtie and then to the human reference genome using TopHat, encompassing some 1.1 billion uniquely mapped reads.

To validate the RNA-seq expression, the researchers used qRT-PCR to examine 15 known immediate-early chlamydia genes, and found a correlation between normalized sequence coverage depth and transcript abundance.

While the number of bacteria-specific transcripts was low — some 0.02 percent of the reads — immediately following infection, those that were expressed in response to infection were generally involved with the chlamydia general secretory pathway, the bacteria's ribosomal subunits, and nutrient uptake. In addition, the researchers reported that a bifunctional riboflavin bioenzyme is expressed early in infection that may mediate the bacteria's uptake of soluble iron from the cell.

Twenty-four hours after infection, a greater proportion of reads — 28.4 percent —emanated from C. trachomatis, which the researchers noted coincided with the bacteria's developmental cycle. While this time point has been better studied than earlier in the infection process, the researchers reported that three ferredoxin-related genes were expressed at that time that had not previously been described.

Meanwhile in the host, infection with C. trachomatis leads to the transcript of a number of immune-response genes like cytokines, chemokines, and signaling molecules. To determine which transcripts were infection-specific, the researchers compared the transcripts from the infected versus mock-infected cells to see which were differentially transcribed.

A portion of those differentially transcribed genes include a number of extracellular matrix components that Myers and his colleagues suspect may affect the scarring associated with chlamydia infection.

For instance, the researchers noted an upregulation of the matrix metalloproteinase MMP-2, increased expression of members of the collagen superfamily — such as COL25A1 — and other basement membrane proteins.

In particular, though, the researchers suggest that dysregulated expression of tenascin C, gremlin1, and TGF-β may be linked to scarring.

In their study, tenascin C was upregulated, and it is not typically found in healthy tissue, but in ones that have been injured. Abnormal upregulation of TNC, they said, could lead to increased matrix deposition and scar formation. In addition, it induces TGF-β, which is itself linked to fibrosis. Further, gremlin, an antagonist of the bone morphogenic protein receptors, is also upregulated, and it, too, can induce TGF-β-induced fibrosis. And, in turn, TGF-β induces GREM1 production.

"It seems that a series of dominos start to fall as soon as Chlamydia infect a cell," Myers said. "Depending on how the individual reacts to that infection, these host responses induce a series of positive feedback loops that ultimately amplify production of the disease-causing scars over time."

The Scan

Driving Malaria-Carrying Mosquitoes Down

Researchers from the UK and Italy have tested a gene drive for mosquitoes to limit the spread of malaria, NPR reports.

Office Space to Lab Space

The New York Times writes that some empty office spaces are transforming into lab spaces.

Prion Pause to Investigate

Science reports that a moratorium on prion research has been imposed at French public research institutions.

Genome Research Papers on Gut Microbe Antibiotic Response, Single-Cell RNA-Seq Clues to Metabolism, More

In Genome Research this week: gut microbial response to antibiotic treatment, approach to gauge metabolic features from single-cell RNA sequencing, and more.