Alexander Fleischmann

The EGF Receptor Provides an Essential Survival Signal for SOS-Dependent Skin Tumor Development

The EGF receptor (EGFR) is required for skin development and is implicated in epithelial tumor formation. Transgenic mice expressing a dominant form of Son of Sevenless (SOS-F) in basal keratinocytes develop skin papillomas with 100% penetrance. However, tumor formation is inhibited in a hypomorphic (wa2) and null EGFR background. Similarly, EGFR-deficient fibroblasts are resistant to transformation by SOS-F and rasV12, however, tumorigenicity is restored by expression of the anti-apoptotic bcl-2 gene. The K5-SOS-F papillomas and primary keratinocytesfrom wa2 mice display increased apoptosis, reduced Akt phosphorylation and grafting experiments imply a cell-autonomous requirement for EGFR in keratinocytes. Therefore, EGFR functions as a survival factor in oncogenic transformation and provides a valuable target for therapeutic intervention in a broader range of tumors than anticipated.

Driving Opposing Behaviors with Ensembles of Piriform Neurons

Anatomic and physiologic studies have suggested a model in which neurons of the piriform cortex receive convergent input from random collections of glomeruli. In this model, odor representations can only be afforded behavioral significance upon experience. We have devised an experimental strategy that permits us to ask whether the activation of an arbitrarily chosen subpopulation of neurons in piriform cortex can elicit different behavioral responses dependent upon learning. Activation of a small subpopulation of piriform neurons expressing channelrhodopsin at multiple loci in the piriform cortex, when paired with reward or shock, elicits either appetitive or aversive behavior. Moreover, we demonstrate that different subpopulations of piriform neurons expressing ChR2 can be discriminated and independently entrained to elicit distinct behaviors. These observations demonstrate that the piriform cortex is sufficient to elicit learned behavioral outputs in the absence of sensory input. These data imply that the piriform does not use spatial order to map odorant identity or behavioral output.

A biophysical signature of network affiliation and sensory processing in mitral cells

One defining characteristic of the mammalian brain is its neuronal diversity. For a given region, substructure, layer or even cell type, variability in neuronal morphology and connectivity persists. Although it is well known that such cellular properties vary considerably according to neuronal type, the substantial biophysical diversity of neurons of the same morphological class is typically averaged out and ignored. Here we show that the amplitude of hyperpolarization-evoked sag of membrane potential recorded in olfactory bulb mitral cells is an emergent, homotypic property of local networks and sensory information processing. Simultaneous whole-cell recordings from pairs of cells show that the amount of hyperpolarization-evoked sag potential and current (Ih) is stereotypic for mitral cells belonging to the same glomerular circuit. This is corroborated by a mosaic, glomerulus-based pattern of expression of the HCN2 (hyperpolarization-activated cyclic nucleotide-gated channel 2) subunit of the Ih channel. Furthermore, inter-glomerular differences in both membrane potential sag and HCN2 protein are diminished when sensory input to glomeruli is genetically and globally altered so that only one type of odorant receptor is universally expressed. Population diversity in this intrinsic property therefore reflects differential expression between local mitral cell networks processing distinct odour-related information.

Rhabdomyosarcoma development in mice lacking Trp53 and Fos: Tumor suppression by the Fos protooncogene

The Fos protein, a major component of the AP-1 transcription factor, is essential for osteoclast differentiation, acts as an oncogene, potentiates transforming signals, and controls invasive growth and angiogenesis during tumor progression. To investigate a potential genetic interaction between the Trp53 and Fos pathways, Trp53/Fos double knockout mice were generated. These mice develop highly proliferative and invasive rhabdomyosarcomas of the facial and orbital regions, with more than 90% penetrance at 6 months of age. Rhabdomyosarcoma cell lines established from the primary tumors express characteristic muscle-specific markers, and reexpression of Fos is associated with enhanced apoptosis in vitro. Moreover, Fos is able to repress Pax7 expression in rhabdomyosarcoma cell lines and primary myoblasts, suggesting a molecular link to genetic alterations involved in human rhabdomyosarcomas.

Olfactory Deficits Cause Anxiety-Like Behaviors in Mice

Anxiety disorders are characterized by persistent fear in the absence of immediate threat and represent the most common psychiatric diseases, with an estimated 28% lifetime prevalence worldwide (Kessler et al., 2010). While symptoms of anxiety are typically evoked by sensory stimuli, it is unknown whether sensory deficits contribute to the development of anxiety disorders. Here we examine the effect of defined genetic mutations that compromise the function of the olfactory system on the development of anxiety-like behaviors in mice. We show that the functional inactivation of the main olfactory epithelium, but not the vomeronasal organ, causes elevated levels of anxiety. Anxiety-like behaviors are also observed in mice with a monoclonal nose, that are able to detect and discriminate odors but in which the patterns of odor-evoked neural activity are perturbed. In these mice, plasma corticosterone levels are elevated, suggesting that olfactory deficits can lead to chronic stress. These results demonstrate a central role for olfactory sensory cues in modulating anxiety in mice.

Fra-1 substitutes for c-Fos in AP-1-mediated signal transduction in retinal apoptosis.

Lack of the AP‐1 member c‐Fos protects photoreceptors against light‐induced apoptosis, a model for retinal degeneration. In mice, light damage increases the activity of the transcription factor AP‐1, while pharmacological suppression of AP‐1 prevents apoptosis, suggesting the involvement of pro‐apoptotic AP‐1 target genes. Recently, however, it was shown that photoreceptors expressing Fra‐1 in place of c‐Fos (FosFosl1/Fosl1) are apoptosis competent despite the lack of transactivation domains in Fra‐1. Here, we show that morphological features of light‐induced apoptosis were indistinguishable in FosFosl1/Fosl1 and wild‐type mice. Furthermore, light exposure comparably increased AP‐1 activity in both genotypes. Opposite to wild‐type mice, Fra‐1, but not c‐Fos, was detectable in AP‐1 complexes of FosFosl1/Fosl1 mice. Importantly, AP‐1 responsiveness for glucocorticoid receptor‐mediated inhibition was preserved in FosFosl1/Fosl1 mice. Thus, Fra‐1 takes over for c‐Fos in pro‐ and anti‐apoptotic signal transduction. As Fra‐1 lacks transactivation domains, AP‐1 may not induce, but rather suppress genes in retinal light damage.

Functional interrogation of an odorant receptor locus reveals multiple axes of transcriptional regulation.

A transgenic approach in mice allows the functional interrogation of an odorant receptor locus in vivo and reveals characteristics of its monogenic and monoallelic expression.

Massive normalization of olfactory bulb output in mice with a 'monoclonal nose'

Perturbations in neural circuits can provide mechanistic understanding of the neural correlates of behavior. In M71 transgenic mice with a “monoclonal nose”, glomerular input patterns in the olfactory bulb are massively perturbed and olfactory behaviors are altered. To gain insights into how olfactory circuits can process such degraded inputs we characterized odor-evoked responses of olfactory bulb mitral cells and interneurons. Surprisingly, calcium imaging experiments reveal that mitral cell responses in M71 transgenic mice are largely normal, highlighting a remarkable capacity of olfactory circuits to normalize sensory input. In vivo whole cell recordings suggest that feedforward inhibition from olfactory bulb periglomerular cells can mediate this signal normalization. Together, our results identify inhibitory circuits in the olfactory bulb as a mechanistic basis for many of the behavioral phenotypes of mice with a “monoclonal nose” and highlight how substantially degraded odor input can be transformed to yield meaningful olfactory bulb output. DOI: http://dx.doi.org/10.7554/eLife.16335.001

Odor identity coding by distributed ensembles of neurons in the mouse olfactory cortex

Olfactory perception and behaviors critically depend on the ability to identify an odor across a wide range of concentrations. Here, we use calcium imaging to determine how odor identity is encoded in olfactory cortex. We find that, despite considerable trial-to-trial variability, odor identity can accurately be decoded from ensembles of co-active neurons that are distributed across piriform cortex without any apparent spatial organization. However, piriform response patterns change substantially over a 100-fold change in odor concentration, apparently degrading the population representation of odor identity. We show that this problem can be resolved by decoding odor identity from a subpopulation of concentration-invariant piriform neurons. These concentration-invariant neurons are overrepresented in piriform cortex but not in olfactory bulb mitral and tufted cells. We therefore propose that distinct perceptual features of odors are encoded in independent subnetworks of neurons in the olfactory cortex. DOI: http://dx.doi.org/10.7554/eLife.26337.001

An Expression Refinement Process Ensures Singular Odorant Receptor Gene Choice

Odorant receptor (OR) gene choice in mammals is a paradigmatic example of monogenic and monoallelic transcriptional selection, in which each olfactory sensory neuron (OSN) chooses to express one OR allele from over 1,000 encoded in the genome [1-3]. This process, critical for generation of the circuit from nose to brain [4-6], is thought to occur in two steps: a slow initial phase that randomly activates a single OR allele, followed by a rapid feedback that halts subsequent expression [7-14]. Inherent in this model is a finite failure rate wherein multiple OR alleles may be activated prior to feedback suppression [15, 16]. Confronted with more than one receptor, the neuron would need to activate a refinement mechanism to eliminate multigenic OR expression and resolve unique neuronal identity [16], critical to the generation of the circuit from nose to olfactory bulb. Here we used a genetic approach in mice to reveal a new facet of OR regulation that corrects adventitious activation of multiple OR alleles, restoring monogenic OR expression and unique neuronal identity. Using the tetM71tg model system, in which the M71 OR is expressed in >95% of mature OSNs and potently suppresses the expression of the endogenous OR repertoire [10], we provide clear evidence of a post-selection refinement (PSR) process that winnows down the number of ORs. We further demonstrate that PSR efficiency is linked to OR expression level, suggesting an underlying competitive process and shedding light on OR gene switching and the fundamental mechanism of singular OR choice.