Fig 2: Manipulating the expression of Zbtb7a in WT and TG2-/- astrocytes differentially impacts morphology. a. Immunoblots showing knockdown (KD) or overexpression (OE) of Zbtb7a, compared to control (CTL), in WT and TG2-/- astrocytes. Since the Zbtb7a OE virus encodes human Zbtb7a, the Zbtb7a bands in these samples migrated slightly less far down the gel than bands in the CTL and KD samples. b. Representative images of transduced astrocyte GFAP network (scale bar = 20 µm) and c. Quantitation of GFAP network area (n=34–50 astrocytes per condition, **p<0.0001), ****p<0.0001).

Fig 3: TG2-/- astrocytes promote neurite outgrowth to a greater extent than WT astrocytes on an inhibitory matrix, a. Representative MAP2 images of neurons grown on PDL (-) or PDL+CSPG (+) matrix and paired with WT or TG2-/- astrocytes (scale bar = 20 µm) b. Quantitation of neurite length on CSPG (n=39–57 neurons per condition, *p<0.05, **p<0.01, ***p<0.001). c. Quantitation of primary neurite number on CSPG (n=43–68 neurons per condition, ***p<0.001).

Fig 4: Knocking down or overexpressing Zbtb7a in WT or TG2-/- differentially impacts neurite outgrowth on a permissive substrate. a. Representative MAP2 images of neurons that were paired with Zbtb7a control (CTL), knockdown (KD) or overexpression (OE) transduced astrocytes (scale bar = 20 µm). b. Quantitation of neurite length on PDL (n=38–49 neurons per condition, ***p<0.001).

Fig 5: Astrocytes facilitate synapse formation among neurons on a permissive, but not inhibitory matrix, independent of the presence of TG2. a. Images of PSD95/Synaptophysin immunostaining of neurons grown on PDL without astrocytes or with WT or TG2-/- astrocytes (scale bar = 20 µm), b. Images of PSD95/synaptophysin immunostaining of neurons grown on CSPG inhibitory matrix without astrocytes or with WT or TG2-/- astrocytes (scale bar = 20 µm), c. Quantification of neuronal synapses approximated by PSD95 puncta colocated with Synaptophysin puncta exclusively in neurites (18–24 neurons per group, ****p<0.001). d. Representative images of Homer/Bassoon immunostaining for neurons grown on PDL and PDL+CSPG (scale bar = 20 µm), and quantification of synapses on PDL approximated by Homer puncta colocated with Bassoon puncta exclusively in neurites (22–23 neurons per group, ***p<0.001).