Gene Expression Profiles of Chlamydophila pneumoniae during the Developmental Cycle and Iron Depletion–Mediated Persistence

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Abstract
The obligate intracellular, gram-negative bacterium Chlamydophila pneumoniae (Cpn) has impact as a human pathogen. Little is known about changes in the Cpn transcriptome during its biphasic developmental cycle (the acute infection) and persistence. The latter stage has been linked to chronic diseases. To analyze Cpn CWL029 gene expression, we designed a pathogen-specific oligo microarray and optimized the extraction method for pathogen RNA. Throughout the acute infection, ratio expression profiles for each gene were generated using 48 h post infection as a reference. Based on these profiles, significantly expressed genes were separated into 12 expression clusters using self-organizing map clustering and manual sorting into the “early”, “mid”, “late”, and “tardy” cluster classes. The latter two were differentiated because the “tardy” class showed steadily increasing expression at the end of the cycle. The transcriptome of the Cpn elementary body (EB) and published EB proteomics data were compared to the cluster profile of the acute infection. We found an intriguing association between “late” genes and genes coding for EB proteins, whereas “tardy” genes were mainly associated with genes coding for EB mRNA. It has been published that iron depletion leads to Cpn persistence. We compared the gene expression profiles during iron depletion–mediated persistence with the expression clusters of the acute infection. This led to the finding that establishment of iron depletion–mediated persistence is more likely a mid-cycle arrest in development rather than a completely distinct gene expression pattern. Here, we describe the Cpn transcriptome during the acute infection, differentiating “late” genes, which correlate to EB proteins, and “tardy” genes, which lead to EB mRNA. Expression profiles during iron mediated–persistence led us to propose the hypothesis that the transcriptomic “clock” is arrested during acute mid-cycle. Chlamydophila (Chlamydia) pneumoniae (Cpn) accounts for approximately one-tenth of the cases of community-acquired pneumonia worldwide, and persistent Cpn infections are thought to be associated with a variety of chronic diseases. Little is known about Cpn transcriptome changes during its biphasic developmental cycle (the acute infection) and persistence stages. Iron limitation, among several other treatments, has recently been shown to lead to persistent Cpn infection. How this pathogen reacts to iron-limiting host defense mechanisms is of great interest, as iron is an important factor affecting virulence. This article reports on the Cpn transcriptome during the developmental cycle and iron depletion–mediated persistence and reveals that genes coding for proteins of the infectious particle (the elementary body [EB]) were expressed constantly at the end of the cycle. In contrast, genes contributing to EB mRNA but not to EB protein showed an increasing expression at the end of the cycle. This suggested that most EB proteins are made in mid-cycle, and the redifferentiation process is initiated only by a limited number of genes. During iron depletion–mediated persistence, the Cpn transcriptome was altered in such a way that an arrest in Cpn gene expression can be proposed.