Fig 1.
De novo 8 bp deletion in the Nbeal2 gene.
The whole exome sequenced G6-ENU mouse inherited the Nbeal2 deletion from a non-ENU parent 31925 (A). Sanger sequencing validates the heterozygous frameshift mutation in the suppressor pedigree (B). Western blot analysis of washed mouse platelets show a band at the expected size for NBEAL2 (~305kDa) in wildtype mice. This band is missing in Nbeal2tm1Lex/tm1Lex mice as well as mice homozygous for the Nbeal2gps allele (C). Schematic overview of the Nbeal2 gene, the location of the deletion and the expected frameshift (D).
Table 1.
Overview of the exonic variants called from whole exome sequencing in 4 mice from the MF5L6 pedigree.
Table 2.
Expected and observed number of progeny in Nbeal2gps/+ crosses.
Fig 2.
Platelet counts are lower in Nbeal2gps/gps mice compared to control mice in both set 1 (A) and set 2 (B) mice while hemoglobin levels are similar between the two groups (C). Nbeal2gps/gps mice from set 1 exhibit significant neutropenia (D), which is not observed in set 2 by CBC (E) or flow cytometry (F). Mean platelet volume (G) and area (H) do not differ in set 1 mice, but show an increase in size for Nbeal2gps/gps mice in set 2 (I).
Table 3.
CBC mean values and standard deviations by each phenotype in set 1.
Fig 3.
Deficiency in platelet alpha granules.
Nbeal2gps/gps platelets appear pale compared to wildtype (black arrows, A). Transmission electron microscopy (TEM) images show dark alpha granules in wildtype platelets (black arrows), which are missing in Nbeal2gps/gps platelets. Red arrows indicate mitochondria (B).
Table 4.
Intensity of platelet staining and frequency of emperipolesis events in bone marrow megakaryocytes.
Fig 4.
Emperipolesis of neutrophils in bone marrow and spleen of NBEAL2 deficient mice.
Increased emperiopolesis of neutrophils (black arrows) in Nbeal2gps/gps mice compared to wildtype was observed in both histologic (A) and cytologic (B) preparations of bone marrow as well as spleen (B and D, respectively).