Fig. 2. Molecular markers and cell position can be used to identify each RP motoneuron in the NB3-1 lineage. (A) RP1 and RP4 are Hb⁺, Kr⁺, Cut^-, Zfh2^- and are shown in insets because they are usually obstructed in the projection (n>100). RP3 is Hb^-, Kr⁺, Cut^-, Zfh2⁺ (n>100). RP5 is Hb^-, Kr^-, Cut⁺, Zfh2⁺ (n>100). A single representative hemisegment of a wild-type (wt) stage-16 CNS is shown as a maximum intensity projection. Midline, left; anterior, up. An expression summary is shown above. (B) (Top row) RP motoneurons are Islet⁺ HB9⁺ (outlined). RP1 and RP4 occupy the deepest layer; RP1 is more dorsal and expresses HB9 at higher levels than does RP4 after stage 15 (n>100). RP3 is directly ventral to RP1 and RP4 (n>100). RP5 is ventral and anterior to RP3 (n>100). White arrowheads, NB7-3-derived EW interneurons. Dashed vertical line, midline. Ventral views of two segments are shown from deep (left) to superficial (right) focal planes. (Bottom row) Each RP neuron has a non-RP neuron sibling, based the duplication of each RP neuron in sanpodo mutants. Eight Islet⁺ HB9⁺ Late Bloomer⁺ RP motoneurons are observed in each hemisegment (quantified in Table 1). Midline, between each pair of panels. (C) Recombination-induced activation of lacZ in NB3-1 labeled RP4, RP3, RP5 and the late-born interneurons but not RP1, showing that RP1 is the first-born neuron in the lineage. HB9, green; β-galactosidase, magenta. RP neurons and NB3-1 are outlined and labeled. One hemisegment of a stage-16 CNS is shown as a maximum intensity projection. Midline, left; anterior, up. A schematic summary of the clone is shown to the right. (D) Schematic of NB3-1 gene expression and cell lineage. The vertical dashed line represents the transition between RP neuron and subsequent interneuron specification. Nb, sibling cell fate specified by Numb; N, sibling cell fate requires Sanpodo and active Notch signaling. Scale bars: 3 µm.