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Analysis of DNA uptake biases in bacteria with and without uptake specificity

University of British Columbia

DNA uptake is the first step in natural transformation of bacteria, leading to DNA internalization and recombination. It is, therefore, a key determinant in genome evolution. Most bacteria take up DNA indiscriminately, but in Pasteurellaceae and Neisseriaceae, the uptake machinery binds preferentially to short sequences enriched in their genome, called DNA uptake signal sequences (USS or DUS). This enrichment is responsible for bacteria preferentially taking up DNA from close relatives, called ‘self-specificity’. My study’s goal was to characterize factors influencing uptake bias across the genome (chapter 2), as well as determined if uptake biases were present in a species without self-specificity (chapter 3). Chapter 2 describes my genome-wide analysis of DNA uptake by Haemophilus influenzae, a species with strong uptake bias, using both measured uptake and the predictions from a simulation model of DNA uptake. Genomic maps of DNA uptake were developed by recovering DNA after it was taken up and deep sequencing it, using DNA preparations sheared into small (50-800 bp) and large (1.5-17 kb) fragment sizes. For short donor DNA fragments, I found a strong uptake bias, up to 1000-fold, that was proportional to the similarity of the USSs to the consensus, including in DNA topology features. I found no evidence of an effect on uptake of non-USS sequences. Uptake of large donor DNA fragments had much less variation than short fragments, with 90% of genomic positions having uptake within 2-fold of the mean. However, it was difficult to assess the USS-dependent factors responsible for this variation because of stochastic noise arising from intrinsic biases of the DNA sequencing process. Chapter 3 describes an analysis to determine if uptake biases are present in Acinetobacter baylyi, a species without self-specificity. This was done by sequencing a degenerate region of synthetic DNA fragments, recovered after uptake by competent A. baylyi cells. Recovered fragments had the same sequence distribution as input DNA, which suggests that there is no evidence of uptake biases.

Text

2021-01-06

Attribution-NonCommercial-NoDerivatives 4.0 International

http://hdl.handle.net/2429/76970

Zoology

University of British Columbia

UBCV

http://creativecommons.org/licenses/by-nc-nd/4.0/