Figure 1.

Functional analysis of CSF3R-T595V mutant in myeloid progenitor cell assay. In vitro colony growth of Csf3r deficient murine hematopoietic progenitor cells expressing the wild-type human CSF3R receptor (wt), the T595I and the T595V mutant, substituting a threonine at amino acid position 595 for an isoleucine or a valine, respectively. CSF3R expressing constructs in pBABE-puro were generated as previously described.¹² The T595V mutation was introduced in pBABE-puro using the QuikChange II XL Site-Directed Mutagenesis Kit (Agilent, Santa Clara, CA, USA). Murine colony assays were performed as previously described.¹² Colonies were grown in the presence of puromycin, either without growth factor (no GF) or with CSF3. The transduction efficiency was corrected for by dividing the number of CSF3-induced colonies by the number of CSF2 (granulocyte macrophage-colony stimulating factor, GM-CSF) induced colonies under puromycin selection as the CSF3R constructs confer puromycin resistance, but do not affect CSF2 induced colony growth. See Online Supplementary Design and Methods and Table S1.