br Author Contributions br Acknowledgments br Introduction N

Author Contributions
Acknowledgments
Introduction Neurons in the CNS are particularly sensitive to injury and degenerative conditions that frequently result in cell death. Although adult neurogenesis persists in restricted 5-aminosalicylic acid areas (Gage, 2000; Hsieh, 2012; Kriegstein and Alvarez-Buylla, 2009; Lie et al., 2004), neurons do not regenerate in most regions of the adult CNS. An unmet challenge in neural injury and degeneration repair is how to replenish lost neurons for functional recovery. Cell fate reprogramming provides new means for regenerating damaged or dead neurons (Arlotta and Berninger, 2014; Cherry and Daley, 2012; Matsui et al., 2014). Not only can cells in culture be reprogrammed into pluripotent stem cells (Takahashi and Yamanaka, 2006), lineage-restricted stem cells (Kim et al., 2011a; Ring et al., 2012), and different postmitotic cell fates (Heinrich et al., 2010, 2011; Karow et al., 2012; Liu et al., 2013; Vierbuchen et al., 2010), but they are also amenable to in vivo fate conversion (Guo et al., 2014; Heinrich et al., 2014; Niu et al., 2013; Qian et al., 2012; Song et al., 2012; Su et al., 2014a, b; Torper et al., 2013; Zhou et al., 2008). In regards to the CNS, resident glial cells have been directly or indirectly converted into functional neurons in the adult brain and spinal cord (Guo et al., 2014; Heinrich et al., 2014; Niu et al., 2013; Su et al., 2014a, b; Torper et al., 2013). Glial cells are broadly distributed and comprise nearly half of the cells in the mammalian CNS. These cells become reactive, proliferate, and form glial scars in response to neural injuries and degeneration (Karimi-Abdolrezaee and Billakanti, 2012; Sofroniew, 2009). These reactive responses are initially beneficial, restricting the spread of damage, but ultimately are deleterious, acting as both a physical and chemical barrier to neuronal regeneration (Karimi-Abdolrezaee and Billakanti, 2012; Sofroniew, 2009). Reprogramming some of these glial cells to functional neurons may constitute a novel therapeutic strategy for diseases associated with the CNS. Through in vivo screens of candidate factors that are able to induce neurogenesis in non-neurogenic regions of the adult brain and spinal cord, we previously showed that the ectopic expression of SOX2 is sufficient to reprogram resident astrocytes to DCX+ induced adult neuroblasts (iANBs) (Niu et al., 2013; Su et al., 2014a). These iANBs pass through a proliferative state and generate mature neurons when supplied with neurotrophic factors. This SOX2-driven in vivo reprogramming process sharply contrasts direct lineage conversion strategies (Guo et al., 2014; Qian et al., 2012; Song et al., 2012; Su et al., 2014b; Torper et al., 2013; Zhou et al., 2008), which change cell fate in a linear fashion without amplification of the induced cell population. However, the cellular mechanism underlying SOX2-dependent in vivo reprogramming of astrocytes was unclear. Furthermore, the subtypes of iANB-derived neurons were not well characterized. In this study, we reveal that SOX2-driven reprogramming of astrocytes transits through intermediate neural progenitor states before the adoption of a mature neuron fate. Immunohistochemistry and electrophysiology further show that induced neurons are functionally mature and predominantly express the marker calretinin.
Results
Discussion This study used genetic fate mapping to reveal that SOX2-driven in vivo reprogramming of adult astrocytes passes through a sequence of distinct cell states (Figure 7), which mimics aspects of endogenous neurogenesis. Ectopic SOX2 converts astrocytes to ASCL1+ intermediate progenitors, which proliferate and generate DCX+ neuroblasts. These early neuroblasts can expand and eventually generate mature neurons when supplied with neurotrophic factors. Expansion through the intermediate progenitors and neuroblasts enable one reprogrammed astrocyte to make multiple functional neurons, which might be therapeutically beneficial for regenerative medicine in treating neural injuries or degeneration.