GATA1-Cre–mediated loxPiga recombination in erythroid cells.

GATA1-Cre–mediated loxPiga recombination leads to primitive and definitive red blood cells that are completely or partially deficient in GPI-linked proteins. (A) Flow cytometric analysis for the expression of CD24 on red blood cells from L^+/− control mice, G^+/−L^+/−, G^+/−L^+/+, and E^+/−L^+/− at birth and at the age of 4 months (4M). E^+/−L^+/− mice are offspring from theEIIa-cre × L breeding and are mosaic for a nonfunctional PIGA protein.8 Note that while a proportion of red cells in E^+/−L^+/− mice is completely deficient in CD24, in GL mice the affected proportion of red cells have residual expression of CD24. Numbers indicate the percentage of cells with normal decreased expression of CD24. (B) Time course of the relative proportion of blood cells deficient in GPI-linked proteins in G^+/−L^+/− and G^+/−L^+/+ mice during an observation period of 16 and 8 months, respectively. Means and SDs are shown. Note that in G^+/−L^+/− mice, the proportion of reticulocytes deficient in CD24 is always higher than the proportion of GPI-anchor–deficient red cells (P ≤ .05). (C) Decreasing levels of residual CD24 expression on the affected red cell population in GL mice during the observation period. The y-axis represents the mean fluorescence intensity of red cells deficient in GPI-linked proteins normalized to the mean fluorescence intensity determined at birth. (D) Expression of CD24 on red cells from 4 representative fetuses at E13.5. At this stage in development, 2 circulating red cell populations can be identified in healthy fetuses by flow cytometry. One population has high side scatter characteristics (SSC^high) and a low level of CD24 expression (CD24^low), which corresponds to circulating primitive red cells. Red cells derived from definitive hematopoiesis are SSC^low and CD24^high (left panel). Note that both E^+/−L^+/− and GL mice have a proportion of primitive and definitive red cells deficient in CD24. However, in contrast to E^+/−L^+/− mice, in whom PIGA⁻ red cells completely lack the expression of CD24,8 the affected primitive and definitive red blood cell populations in GL mice show a wide distribution of residual expression of CD24. SSC indicates side scatter; L^+/−, healthy control mouse. E^+/−L^+/− mice are offspring from the EIIa-cre × L breeding.8 Numbers indicate the percentage of cells with normal decreased expression of CD24.