Figure 1
Expression of the endogenous and transgenic SEPT4 in the mouse brain. (A) Representative parasagittal brain sections of the wild-type (wt, Sept4+/+) and transgenic (tg, Sept4Tg/+) adult male mice stained with hematoxylin and eosin (HE, left panels), or immunostained for SEPT4 (right panels). In Sept4Tg/+ brain, the increment of SEPT4 is the most obvious where the endogenous SEPT4 expression is relatively low (e.g., the striatum, hippocampus and cerebral cortex). No recognizable anatomical and histological difference was found between Sept4+/+ and Sept4Tg/+ brains. (B) Quantitative immunoblot analysis of the major SEPT4 polypeptides in fractionated lysates from Sept4+/+ and Sept4Tg/+ brain regions. (Top) Representative immunoblot patterns for SEPT4 and α-tubulin (loading control) of the following brain regions; the cerebral cortex (cx), striatum (str), midbrain and brainstem (mb + bs), and cerebellum (cbl). The lysate (containing 3 μg protein) from each brain region was separated by ultracentrifugation into the supernatant/soluble (s) fraction and the pellet/insoluble (p) fraction. Immunoblot with the SEPT4 antibody (used in panel A) consistently detected four major bands of approximately 54, 52, 48 and 44 kDa, all of which were absent in Sept4−/− mice ([12]; data not shown). The top band, containing the transgene-encoded 54 kDa isoform was consistently increased in each region of Sept4Tg/+ brain, reflecting the pan-neural expression by the prion promoter. (Bottom) Comparison of the total densitometric values of the four major SEPT4 polypeptides in the two fractions of the brain regions. The ratio of soluble SEPT4 content of Sept4Tg/+ brain over Sept4+/+ brain was roughly x 1.5, except for the cerebellum.