We have investigated two clonal mouse olfactory placode (OP) cell lines as a model system for studying endogenous odorant receptor (OR) regulation. Both lines can be differentiated into bipolar neurons with transcriptional profiles consistent with mature sensory neurons. We show that single cells exhibit monogenic OR expression like sensory neurons in vivo. Monogenic OR expression is established in undifferentiated cells and persists through differentiation, but OR gene choice is not a clonal property of either cell line. Interestingly, OR RNA shifts from predominantly nuclear to cytoplasma during differentiation of both cell lines. Finally, our data indicate that a restricted subset of OR genes and OR clusters are over-represented in cell populations, suggesting either a pre-existing intrinsic bias in OP founder cells or extrinsic influences arising from culture conditions. Keywords: differentiation, odorant receptor, regulation, transcription. J. Neurochem. (2009) 108, 486-497.JOURNAL OF NEUROCHEMISTRY | 2009NEUROCHEMISTRY | | 108 | 486-497 doi: 10.1111NEUROCHEMISTRY | /j.1471NEUROCHEMISTRY | -4159.2008 Production of a functional OR protein is required to ensure the 'locking in' of the selected OR gene (Serizawa et al. 2003;Lewcock and Reed 2004). If a sensory neuron initially transcribes an OR gene that does not encode an intact open reading frame, and thus cannot be translated into a functional OR protein, the sensory neuron will make another OR choice. Feedback regulation ensures that the sensory neuron will be productive, as the genome is populated with OR pseudogenes that might not have yet accumulated enough mutations to render their promoters non-functional. Thus, the process leading to monogenic OR expression seems to involve three discrete steps: specification of a subset of ORs (e.g., ORs appropriate for a spatial zone in the olfactory epithelium), random selection of one OR among specified subsets, and commitment to the selected OR via feedback regulation.The existence of a suitable OSN cell line could greatly facilitate investigation of the molecular mechanisms of OR regulation. Several cell lines derived from olfactory tissue have been described (Largent et al. 1993;MacDonald et al. 1996;Goldstein et al. 1997;Murrell and Hunter 1999;Barber et al. 2000;Illing et al. 2002), yet to our knowledge, only two of these lines appear to express endogenous ORs (Illing et al. 2002). These two clonal olfactory placode (OP) cell lines, termed OP6 and OP27, were characterized by gene expression profiling as derived from intermediate-late stages in the OSN cell lineage. Both lines are immortalized by a temperature-sensitive large T-antigen transgene, and can be induced by retinoic acid at the non-permissive temperature to differentiate into bipolar neurons that express markers for OSNs, including OR genes. In previous studies, only one OR gene, Olfr1355, was identified in differentiated OP27 cell populations, and two closely related OR genes, Olfr57 and Olfr1351, were identified in differentiated OP6...

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1. All giant interneurons (GIs) were ablated from the nerve cord of cockroaches by electrocautery, and escape behavior was analyzed with high-speed videography. Animals with ablations retained the ability to produce wind-triggered escape, although response latency was increased (Table 1, Fig. 4). Subsequent lesions suggested that these non-GI responses depended in part on receptors associated with the antennae.2. Antennal and cercal systems were compared by analyzing escape responses after amputating either cerci or antennae. With standard wind stimuli (high peak velocity) animals responded after either lesion. With lower intensity winds, animals lost their ability to respond after cercal removal (Fig. 6).3. Removal of antennae did not cause significant changes in behavioral latency, but in the absence of cerci, animals responded at longer latencies than normal (Fig. 7).4. The cercal-to-GI system can mediate short latency responses to high or low intensity winds, while the antennal system is responsive to high intensity winds only and operates at relatively longer latencies. These conclusions drawn from lesioned animals were confirmed in intact animals with restricted wind targeting the cerci or antennae only (Fig. 9).5. The antennae do not represent a primary wind-sensory system, but may have a direct mechanosensory role in escape.

G-protein-coupled receptor (GPCR)-kinase interacting protein-1 (GIT1) is a multi-function scaffold protein. However, little is known about its physiological role in the heart. Here we sought to identify the cardiac function of GIT1. Global GIT1 knockout (KO) mice were generated and exhibited significant cardiac hypertrophy that progressed to heart failure. Electron microscopy revealed that the hearts of GIT1 KO mice demonstrated significant morphological abnormities in mitochondria, including decreased mitochondrial volume density, cristae density and increased vacuoles. Moreover, mitochondrial biogenesis-related gene peroxisome proliferator-activated receptor γ (PPARγ) co-activator-1α (PGC-1α), PGC-1β, mitochondrial transcription factor A (Tfam) expression, and total mitochondrial DNA were remarkably decreased in hearts of GIT1 KO mice. These animals also had impaired mitochondrial function, as evidenced by reduced ATP production and dissipated mitochondrial membrane potential (Ψm) in adult cardiomyocytes. Concordant with these mitochondrial observations, GIT1 KO mice showed enhanced cardiomyocyte apoptosis and cardiac dysfunction. In conclusion, our findings identify GIT1 as a new regulator of mitochondrial biogenesis and function, which is necessary for postnatal cardiac maturation.

Erythropoiesis is a highly regulated process that generates enucleate red blood cells from committed erythroid progenitors. Chromatin condensation culminating in enucleation is a defining feature of this process. Setd8 is the sole enzyme that can mono-methylate histone H4, lysine 20 and is highly expressed in erythroblasts compared to most other cell types. Erythroid Setd8 deletion results in embryonic lethality from severe anemia due to impaired erythroblast survival and proliferation. Setd8 protein levels are also uniquely regulated in erythroblasts, suggesting a cell-type-specific role for Setd8 during terminal maturation. Consistent with this hypothesis, Setd8 Δ/Δ erythroblasts have profound defects in transcriptional repression, chromatin condensation, and heterochromatin accumulation. Together, these results suggest that Setd8, used by most cells to promote mitotic chromatin condensation, is an essential aspect of the transcriptional repression and chromatin condensation that are hallmarks of terminal erythroid maturation.