Conditional <i>Klf6</i> inactivation in vivo causes selective loss of differentiating oligodendrocytes.

<p><b>(A–F)</b> Confocal and morphometric analysis of oligodendrocyte numbers in developing <i>Olig1Cre</i>:<i>Klf6</i>^<i>fl/fl</i> and control <i>Klf6</i>^<i>fl/fl</i> spinal cords. Panel <b>(A)</b> shows samples from E12.5, immunostained for Olig2 and either Mnr (upper panels) or NeuN (lower panels). At E12.5, Olig2⁺ numbers in <i>Olig1Cre</i>:<i>Klf6</i>^<i>fl/fl</i> and control samples are identical, and no differences are seen in neuronal markers. In controls, Olig2⁺ cell numbers then increase from E16.5 through P14, but no increase is seen in <i>Olig1Cre</i>:<i>Klf6</i>^<i>fl/fl</i> samples <b>(B–D)</b>. Areas outlined in panel <b>(B)</b> are shown at higher magnification, inset. See also <a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.1002467#pbio.1002467.s006" target="_blank">S4B Fig</a>. At P1, myelin proteins are expressed in controls, but not in <i>Olig1Cre</i>:<i>Klf6</i>^<i>fl/fl</i> spinal cords <b>(C)</b>. <b>(E,F)</b> Analysis of stage-specific markers. Early events in differentiation, such as Nkx2.2 co-expression at E14.5, occur normally in <i>Olig1Cre</i>:<i>Klf6</i>^<i>fl/fl</i> mice, but differentiating cells are selectively lost at subsequent timepoints <b>(E)</b>. In contrast, numbers of Olig2⁺Sox9⁺ OLP remain identical to controls <b>(F)</b>. There are no changes in Mnr⁺ motor neurons, which share the same origin as ventral OLP in the pMN domain (<b>A</b> upper panels, <b>B, G</b>). <b>(H,I)</b> Selective loss of differentiating cells is associated with increased apoptosis <b>(H)</b>, but OLP proliferation is unaffected <b>(I)</b>. <b>(J–L)</b> Postnatal stage-specific analysis confirms absence of differentiating (Apc⁺) oligodendrocytes from <i>Olig1Cre</i>:<i>Klf6</i>^<i>fl/fl</i> mice <b>(J,L)</b>, whereas OLP (Olig2⁺Ng2⁺) numbers are comparable to controls <b>(K,L)</b>. Representative cells are arrowed. <b>(M)</b> Analysis of <i>NG2creER</i>^<i>–</i>:<i>Klf6</i>^<i>fl/fl</i> mice, in which <i>Klf6</i> inactivation is inducibly targeted to OLP. See also <a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.1002467#pbio.1002467.s006" target="_blank">S4C and S4D Fig</a>. Similar to <i>Olig1Cre</i>:<i>Klf6</i>^<i>fl/fl</i> embryos, these mice also display selective loss of differentiating oligodendrocytes. Data are mean ± SEM. Statistics, <b>(D–I,L)</b> ANOVA plus Bonferroni post test, <b>(M)</b> Student’s <i>t</i> test, <i>***p <</i> 0.001. Scale: <b>(A,B)</b> 100 μm, inset 20 μm, <b>(C)</b> 250 μm, <b>(J,K)</b> 10 μm. Data shown are from lumbar sections of two to six mice per genotype per timepoint. Thoracic sections showed compatible findings. Individual values are in <a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.1002467#pbio.1002467.s002" target="_blank">S1 Data</a>.