A NNOTATION OF GLIAL DRIVERS

functional studies without applying elaborate intersectional strategies (del Valle Rodriguez et al., 2012; Venken et al., 2011).

Drivers with unknown pattern were tested for nuclearly expressed GFP in glia using double-labelings with glial-specific REPO antibodies.

5.1 Glial Gal4 driver lines within the Janelia Farm Collection

Among the ~6.650 imaged Gal4 driver lines within the Janelia Farm Collection, we identified 669 with expression in glia (Figure 8A). All glial driver lines were categorized into the 5 major generic glial subtypes – perineurial glia, subperineurial glia, cortex glia, ensheathing glia and astrocyte-like glia (Figure 8). Driver lines with expression in multiple subtypes as well as driver lines with expression in trachea were categorized separately.

Figure 8: Glia in the Janelia Farm Gal4 collection.

A Ten percent of all the driver lines within the JFRC Gal4 Collection contain glial expression. B They can be subdivided into perineurial, subperineurial, cortex, astrocyte-like and ensheathing glia. The number of driver lines specifically expressed in either of these lines is indicated with numbers in the pie chart. Additionally, driver lines, which express in more than one subtype, are registered as Multiple.

Among all generic glial subtypes, perineurial glia drivers are found most frequently (33%), followed by subperineurial glia (14%), ensheathing glia (14%)

ANNOTATION OF GLIAL DRIVERS  

and astrocyte-like glia (12%). Cortex glial drivers are most rare (6%). Expression in more than one generic subtype is observed in 12 percent of all drivers (Figure 8B). Further subdivision placed them into categories according to lines, which express in all glia of one or more generic glial subtypes, All, or only in a region-restricted manner, region-specific. Additionally, we annotated if the driver lines shows

mosaic or non-mosaic expression (Figure 9).

Figure 9: Classification of glial Gal4 drivers into generic subtypes.

Driver lines found within the Janelia Farm Gal4 Collection were categorized according to their global expression characteristics. A Among the glial driver lines, 16 percent express in all glia of one or multiple generic glial subtypes in a non-mosaic fashion (All / non-mosaic); 22 percent express in all glia of one or

 ANNOTATION OF GLIAL DRIVERS  

multiple generic glial subtypes in a mosaic fashion (All / mosaic); 23 percent express in a region specific subset of one or multiple generic glial subtypes in a non-mosaic fashion (Region-specific / non-mosaic); 35 percent express in a region-specific subset of one or multiple generic glial subtypes in a mosaic fashion (Region-specific / mosaic). For further details please refer to the annotation details in the attached excel- spreadsheet. B-F As introduced in Figure 8, subtypes (in this case, all the drivers of one generic subclass) can be subdivided according to expression in the entire subtype or only a particular region, as well as by the degree of mosaicism. The numbers (green) in the figure legend box of each single generic glial subgroup (B-F) depict the number of actual driver lines with corresponding expression pattern. We found 44 driver lines with expression in Trachea. Those drivers are not considered in the present annotation analysis.

Within the glia-containing drivers of the Janelia Farm Gal4 collection, 61 percent show region specific expression pattern; among those, the majority (65 percent) of drivers induce mosaic expression. The minority of drivers promotes expression in all glia of one or multiple generic glial subtypes; among those, the majority (58 percent) induces mosaic expression (Figure 9A).

More specifically, we observe the following subtype characteristics: In the perineurial glial subtype (Figure 9B), the distribution between the different subcategories (All or region-specific / mosaic or non-mosaic) is relatively even, with a drift towards mosaic expression pattern (63%). In the subperineurial glial subtype (Figure 9C), the distribution between the different subcategories is relatively even; among all subgroups, subperineurial glia show the least percentage of region-specific mosaic driver lines. In the cortex glia subtype (Figure 9D), the distribution between the different subcategories is uneven; region- specificity is observed in 75percent of all driver lines with the majority (~2/3) showing mosaic expression patterns. In the astrocyte-like glia subtype (Figure 9E), the distribution between the different subcategories is uneven; mosaicism is observed in 75 percent of all driver lines, half of them show region-specific expression patterns. In the ensheathing glia subtype (Figure 9F), the distribution between the different subcategories is uneven; region-specificity is observed in 75 percent of all driver lines, half of them show mosaic expression patterns. Notably, a driver line that expresses in all ensheathing glia could not be identified; the 3 drivers counted here show high neuronal background or very weak glial expression.

ANNOTATION OF GLIAL DRIVERS  

For the reasons described in the beginning of this section, all glial drivers, after the initial characterization described above, were classified into A, B and C-drivers according to their quality and usability for further analysis (Figure 10A). The results as well as the proportion of generic glial subtypes in the group of A-drivers are presented in (Figure 10B).

Figure 10: Triage of glial driver lines and classification of A-listed drivers into generic subtypes.

A Glial driver found in the Janelia Farm Collection were categorized as introduced previously into A, B and C-lines. B A-listed driver lines were further subdivided into the 5 major generic subclasses and those that label multiple generic subclasses. Since A-listed lines were selected subjectively according to uniqueness of pattern, strength of the driver and usability for functional analysis of glial cell function, global expression characteristics as in Figure 9B-F will not be explicitly plotted here. However, this data is contained Excel- spreadsheet attached to this thesis.

316 lines were A-, 270 lines B- and 127 lines C-‘listed’. Among the A-drivers, we identified 79 with perineurial and 47 with subperineurial glial expression, 23 with cortex glial expression, 48 with astrocyte-like and 58 with ensheathing glial expression. In 46 lines, expression was found in more than one glial subtype (Figure 10B).

 ANNOTATION OF GLIAL DRIVERS  

Altogether, the driver lines contained in the A-list provide us with the necessary bandwidth of drivers to characterize both generic (also see 5.2) and region- specific glial populations (also see 5.3).

ANNOTATION OF GLIAL DRIVERS