SUPPLEMENTAL FIGURE LEGENDS
Supplemental Table 1. siRNA and primer sequences used to generate in situ hybridization
probes and in RT-PCR studies.
Supplemental Table 2. A list of primary and secondary antibodies used in this study.
Supplemental Figure 1. BEHAB/brevican knockdown does not alter self-renewal of glioma
initiating cells in vitro. The ability of isolated GIC clones expressing either control (pLL3.7) or
two independent shB/b contructs (shB/b 1 and shB/b 2) to give rise to spheres was
assessed. A, The percentage of single isolated clones that give rise to spheres was not
statistically significantly different between control and shB/b expressing GICs. B,
Quantification of sphere circumference, as a measure of size, was also not significantly
different.
Supplemental Figure 2. BEHAB/brevican does not alter proneural gene expression in
GICs. Quantitative RT-PCR was done to examine the expression of proneural genes BCAN,
OLIG2, DLL3, ASCL1 following B/b knockdown at 5 DIV after infection with two independent
shRNA contructs (shB/b 1 and shB/b 2) in 0627 GICs. The fold change in gene expression
relative to control levels is presented following normalization to loading controls. The
expression of BCAN was significantly decreased relative to controls, while the expression of
other proneural genes were not significantly altered.
Supplemental Figure 3. BEHAB/brevican knockdown does not alter the ability of glioma
initiating cells to form intracranial grafts. Control (pLL3.7) or B/b knockdown (shB/b) GICs
were injected into the striatum of animal models and tissue was collected at 20 DPI to
assess intracranial graft formation. GFP expression by control or shB/b expressing plasmids
was used to demarcate engrafted GICs. A, Low mag representative image shows control
engrafted GICs localized primarily along the injection tract and diffused in grey matter and
surrounding white matter tracts, scale (1mM). B, C, High magnification images show lateral
invasion of control GICs into grey matter at the tumor/stromal boundary, scale (100μM). D,
E, F, shB/b-GICs form intracranial grafts with lateral invasion similar to controls. In all cases,
nuclei were counterstained with Hoechst (Blue).