Journal: Cell

Article Title: Microtubule-induced Pins/Galphai cortical polarity in Drosophila neuroblasts.

doi: 10.1016/j.cell.2005.09.043

Figure Lengend Snippet: Figure 3. Dlg GK Domain Is Required to Maintain Dlg-Microtubule Alignment (A–E) Lateral view of stage 10 metaphase neuroblasts. Genotypes are listed above panels. Top row, Pins staining with or without PH3. Neuroblasts in (B)–(E) are from the pool represented by solid gray/black bars in Figure 2F (i.e., the minority of dlg insc neuroblasts that form Pins/Gai crescents). Second row, same neuroblast with a-tubulin staining to show spindle position. Third row, double-labeled neuroblasts to show position of the mitotic spindle relative to cortical polarity. White arrowheads indicate center of the crescent; yellow arrows point to spindle poles, along the long axis of the mitotic spindle. Bottom row, quantitation of spindle orientation relative to Pins/Gai crescents. Each red line indicates the angle between the spindle axis and the center of the Pins/ Gai crescent for each metaphase neuroblast scored, represented as a dashed arc in (C) and (E).

Article Snippet: Primary antibodies used for these studies include rabbit phospho-Histone H3 (1:1,000; Upstate), mouse a-tubulin (1:2,000; Sigma-Aldrich), rat a-tubulin (1:100; Serotec), mouse g-tubulin (1:2,000; Sigma-Aldrich), rabbit Pins (Yu et al., 2000), rat Pins (1:500; F. Yu), rabbit Gai (Schaefer et al., 2001), mouse Dlg 4F3E2 (Parnas et al., 2001), rabbit Insc (1:500; W. Chia), rabbit anti-Baz (1:500; A. Wodarz), rabbit aPKC (1:500; Santa Cruz), rat Par-6 (Rolls et al., 2003), and guinea pig Senseless (Nolo et al., 2000).

Techniques: Staining, Labeling, Quantitation Assay

Journal: Cell

Article Title: Microtubule-induced Pins/Galphai cortical polarity in Drosophila neuroblasts.

doi: 10.1016/j.cell.2005.09.043

Figure Lengend Snippet: Figure 5. HA:Khc-73 Localizes to Microtubule Plus Ends in Mitotic Neuroblasts (A) 1.0 to 1.2 mm serial optical sections from a Taxol-treated larval neuroblast from an animal expressing UAS-HA:Khc-73FL induced by worniu-GAL4. Top row, HA staining marking localization of HA:Khc-73FL. Bottom row, same neuroblast triple-labeled with HA, a-tubulin, and PH3. White arrowheads indicate localization of HA:Khc-73FL to microtubule plus ends. Yellow arrowheads indicate localization of HA:Khc-73FL to microtubule plus ends associated with DNA. Scale bar = 5 mm. (B) Projection from the 3D reconstruction of the neuroblast shown in (A) and in Movie S1. The central red puncta is at the plus end of a microtubule projecting toward the reader. Scale bar = 5 mm. (C) As a control, a single optical section from a wild-type Taxol-treated neuroblast showing no background HA immunostaining.

Article Snippet: Primary antibodies used for these studies include rabbit phospho-Histone H3 (1:1,000; Upstate), mouse a-tubulin (1:2,000; Sigma-Aldrich), rat a-tubulin (1:100; Serotec), mouse g-tubulin (1:2,000; Sigma-Aldrich), rabbit Pins (Yu et al., 2000), rat Pins (1:500; F. Yu), rabbit Gai (Schaefer et al., 2001), mouse Dlg 4F3E2 (Parnas et al., 2001), rabbit Insc (1:500; W. Chia), rabbit anti-Baz (1:500; A. Wodarz), rabbit aPKC (1:500; Santa Cruz), rat Par-6 (Rolls et al., 2003), and guinea pig Senseless (Nolo et al., 2000).

Techniques: Expressing, Staining, Labeling, Control, Immunostaining

Journal: Cell

Article Title: Microtubule-induced Pins/Galphai cortical polarity in Drosophila neuroblasts.

doi: 10.1016/j.cell.2005.09.043

Figure Lengend Snippet: Figure 6. Khc-73, Dlg, Pins, and Gai Are Required for Tight Coupling of the Apical Spindle Pole with Cortical Insc/Par Proteins Metaphase neuroblasts scored for apical spindle pole alignment with the aPKC cortical crescent. Markers and genotypes indicated. First row, neuroblasts triple-labeled for aPKC (marked with arrowhead), g-tubulin (marked with yellow arrows), and DNA. Dashed arc indicates distance between the two. Second row, neuroblasts labeled with aPKC (crescents marked with arrowhead) and the DNA marker PH3. Third row, same neuroblast with a-tubulin staining to show position of the mitotic spindle; spindle poles labeled with yellow arrows. Fourth row, same neuroblasts triple-labeled to show position of the mitotic spindle relative to cortical polarity. Bottom row, quantitation of spindle orientation defects. Each red line indicates the angle (represented by the dashed lines in the fourth row panels) between the spindle axis and the center of the Pins/Gai crescent. Only a-tubulin-labeled neuroblasts with a clear apical spindle pole were scored; in some neuroblasts, spindle morphology defects precluded scoring. Neuroblasts are shown at the stage where the phenotype is most pen- etrant (this varies because there are different amounts of residual maternal protein present in each genotype) or at the stage where we have the strongest Gal4 driver. (A, C, and E) Third larval instar. (B) Embryonic stage 10. (D) Second larval instar. Scale bar in top left panel = 5 mm, and scale bar in panel below = 10 mm.

Article Snippet: Primary antibodies used for these studies include rabbit phospho-Histone H3 (1:1,000; Upstate), mouse a-tubulin (1:2,000; Sigma-Aldrich), rat a-tubulin (1:100; Serotec), mouse g-tubulin (1:2,000; Sigma-Aldrich), rabbit Pins (Yu et al., 2000), rat Pins (1:500; F. Yu), rabbit Gai (Schaefer et al., 2001), mouse Dlg 4F3E2 (Parnas et al., 2001), rabbit Insc (1:500; W. Chia), rabbit anti-Baz (1:500; A. Wodarz), rabbit aPKC (1:500; Santa Cruz), rat Par-6 (Rolls et al., 2003), and guinea pig Senseless (Nolo et al., 2000).

Techniques: Labeling, Marker, Staining, Quantitation Assay

Journal: PLoS ONE

Article Title: Characterization of Cognitive Deficits in Rats Overexpressing Human Alpha-Synuclein in the Ventral Tegmental Area and Medial Septum Using Recombinant Adeno-Associated Viral Vectors

doi: 10.1371/journal.pone.0064844

Figure Lengend Snippet: Expression of the transgenic a-syn protein was observed in the MS/vDBB as well as throughout the VTA with negligible expression in the SN (A). High-power photomicrographs reveal human a-syn-immunopositive neurons in the MS/vDBB (B) and the VTA (C). Labeled fibers were seen in the prefrontal cortex (D), nucleus accumbens (E), olfactory tubercule (F), medial striatum (G), and hippocampus (H). In addition, human a-syn positive inclusions were seen in the PFC, NAcc, MS and hippocampus (arrow heads in D, E, G, H). Small beaded structures positive for a-syn were also seen along the fibers in the prefrontal cortex (D). PK (10 µm/ml) digestion of hippocampal sections followed by immunostaining specific for human a-syn (clone syn 211) revealed insoluble human a-syn positive aggregates, regardless of incubation time with PK (I–M). Control sections were stained for an antibody that recognizes endogenous rat a-syn (clone EP1646Y) (N–R). Endogenous a-syn expressed in the CA3 region was completely digested after 90 min of incubation with PK (Q–R). Scale bar indicates 2 mm (A), 30 µm (H), 50 µm (M) and 100 µm (R). Abbreviations: a-syn, alpha-synuclein; Hipp, hippocampus; MS, medial striatum; MS/vDBB, medial septum/vertical limb of the diagonal band of Broca; NAcc, nucleus accumbens; OT, olfactory tubercule; PFC, prefrontal cortex; PK, proteinase K; rAAV5, recombinant adeno associated vectorserotype 5; VTA, ventral tegmental area.

Article Snippet: Sections obtained from a-syn injected rats were incubated with the human specific mouse anti-a-syn antibody (1∶20000; syn 211, 36-008, Millipore) and control sections obtained from non-injected rats were incubated with a rabbit anti-a-syn antibody that recognizes endogenous rat a-syn (1∶1000; EP1646Y, ab51252, Abcam).

Techniques: Expressing, Transgenic Assay, Labeling, Immunostaining, Incubation, Staining, Recombinant