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Cas9+ conditionally-immortalized macrophages as a tool for bacterial pathogenesis and beyond.

ABSTRACT: Macrophages play critical roles in immunity, development, tissue repair, and cancer, but studies of their function have been hampered by poorly-differentiated tumor cell lines and genetically-intractable primary cells. Here we report a facile system for genome editing in non-transformed macrophages by differentiating ER-Hoxb8 myeloid progenitors from Cas9-expressing transgenic mice. These conditionally immortalized macrophages (CIMs) retain characteristics of primary macrophages derived from the bone marrow yet allow for easy genetic manipulation and a virtually unlimited supply of cells. We demonstrate the utility of this system for dissection of host genetics during intracellular bacterial infection using two important human pathogens: Listeria monocytogenes and Mycobacterium tuberculosis.

SUBMITTER: Roberts AW

PROVIDER: S-EPMC6579556 | biostudies-literature | 2019 Jun

REPOSITORIES: biostudies-literature

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Cas9<sup>+</sup> conditionally-immortalized macrophages as a tool for bacterial pathogenesis and beyond.

Roberts Allison W AW Popov Lauren M LM Mitchell Gabriel G Ching Krystal L KL Licht Daniel J DJ Golovkine Guillaume G Barton Gregory M GM Cox Jeffery S JS

eLife 20190617

Macrophages play critical roles in immunity, development, tissue repair, and cancer, but studies of their function have been hampered by poorly-differentiated tumor cell lines and genetically-intractable primary cells. Here we report a facile system for genome editing in non-transformed macrophages by differentiating ER-Hoxb8 myeloid progenitors from Cas9-expressing transgenic mice. These conditionally immortalized macrophages (CIMs) retain characteristics of primary macrophages derived from the ...[more]

PMID: 31204998

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Project description:Redundancy has been referred to as a state of no longer being needed or useful. Microbiologists often theorize that the only case of true redundancy in a haploid organism would be a recent gene duplication event, prior to divergence through selective pressure. However, a growing number of examples exist where an organism encodes two genes that appear to perform the same function. For example, many pathogens translocate multiple effector proteins into hosts. While disruption of individual effector genes does not result in a discernable phenotype, deleting genes in combination impairs pathogenesis: this has been described as redundancy. In many cases, this apparent redundancy could be due to limitations of laboratory models of pathogenesis that do not fully recapitulate the disease process. Alternatively, it is possible that the selective advantage achieved by this perceived redundancy is too subtle to be measured in the laboratory. Moreover, there are numerous possibilities for different types of redundancy. The most common and recognized form of redundancy is functional redundancy whereby two proteins have similar biochemical activities and substrate specificities allowing each one to compensate in the absence of the other. However, redundancy can also exist between seemingly unrelated proteins that manipulate the same or complementary host cell pathways. In this article, we outline 5 types of redundancy in pathogenesis: molecular, target, pathway, cellular process, and system redundancy that incorporate the biochemical activities, the host target specificities and the impact of effector function on the pathways and cellular process they modulate. For each type of redundancy, we provide examples from Legionella pathogenesis as this organism employs over 300 secreted virulence proteins and loss of individual proteins rarely impacts intracellular growth. We also discuss selective pressures that drive the maintenance of redundant mechanisms, the current methods used to resolve redundancy and features that distinguish between redundant and non-redundant virulence mechanisms.

Dataset Information

Cas9+ conditionally-immortalized macrophages as a tool for bacterial pathogenesis and beyond.

Publications

Cas9<sup>+</sup> conditionally-immortalized macrophages as a tool for bacterial pathogenesis and beyond.

Similar Datasets

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