TY - JOUR
T1 - Reverse genetic screen reveals that Il34 facilitates yolk sac macrophage distribution and seeding of the brain
AU - Kuil, Laura E.
AU - Oosterhof, Nynke
AU - Geurts, Samuël
AU - Van Der Linde, Herma C.
AU - Meijering, Erik
AU - Van Ham, Tjakko J.
PY - 2019
Y1 - 2019
N2 - Microglia are brain-resident macrophages, which have specialized functions important in brain development and in disease. They colonize the brain in early embryonic stages, but few factors that drive the migration of yolk sac macrophages (YSMs) into the embryonic brain, or regulate their acquisition of specialized properties, are currently known. Here, we present a CRISPR/Cas9-based in vivo reverse genetic screening pipeline to identify new microglia regulators using zebrafish. Zebrafish larvae are particularly suitable due to their external development, transparency and conserved microglia features. We targeted putative microglia regulators, by Cas9/gRNA complex injections, followed by Neutral-Red-based visualization of microglia. Microglia were quantified automatically in 3-day-old larvae using a software tool we called SpotNGlia. We identified that loss of zebrafish colony-stimulating factor 1 receptor (Csf1r) ligand, Il34, caused reduced microglia numbers. Previous studies on the role of IL34 in microglia development in vivo were ambiguous. Our data, and a concurrent paper, show that, in zebrafish, il34 is required during the earliest seeding of the brain by microglia. Our data also indicate that Il34 is required for YSM distribution to other organs. Disruption of the other Csf1r ligand, Csf1, did not reduce microglia numbers in mutants, whereas overexpression increased the number of microglia. This shows that Csf1 can influence microglia numbers, but might not be essential for the early seeding of the brain. In all, we identified il34 as a modifier of microglia colonization, by affecting distribution of YSMs to target organs, validating our reverse genetic screening pipeline in zebrafish.
AB - Microglia are brain-resident macrophages, which have specialized functions important in brain development and in disease. They colonize the brain in early embryonic stages, but few factors that drive the migration of yolk sac macrophages (YSMs) into the embryonic brain, or regulate their acquisition of specialized properties, are currently known. Here, we present a CRISPR/Cas9-based in vivo reverse genetic screening pipeline to identify new microglia regulators using zebrafish. Zebrafish larvae are particularly suitable due to their external development, transparency and conserved microglia features. We targeted putative microglia regulators, by Cas9/gRNA complex injections, followed by Neutral-Red-based visualization of microglia. Microglia were quantified automatically in 3-day-old larvae using a software tool we called SpotNGlia. We identified that loss of zebrafish colony-stimulating factor 1 receptor (Csf1r) ligand, Il34, caused reduced microglia numbers. Previous studies on the role of IL34 in microglia development in vivo were ambiguous. Our data, and a concurrent paper, show that, in zebrafish, il34 is required during the earliest seeding of the brain by microglia. Our data also indicate that Il34 is required for YSM distribution to other organs. Disruption of the other Csf1r ligand, Csf1, did not reduce microglia numbers in mutants, whereas overexpression increased the number of microglia. This shows that Csf1 can influence microglia numbers, but might not be essential for the early seeding of the brain. In all, we identified il34 as a modifier of microglia colonization, by affecting distribution of YSMs to target organs, validating our reverse genetic screening pipeline in zebrafish.
KW - Brain development
KW - Hematopoiesis
KW - Macrophages
KW - Microglia
KW - Reverse genetic screen
UR - http://www.scopus.com/inward/record.url?scp=85062875365&partnerID=8YFLogxK
U2 - 10.1242/dmm.037762
DO - 10.1242/dmm.037762
M3 - Article
C2 - 30765415
SN - 1754-8403
VL - 12
JO - DMM Disease Models and Mechanisms
JF - DMM Disease Models and Mechanisms
IS - 3
M1 - dmm037762
ER -