Faculty Bibliography

Stochastic mechanisms are sometimes utilized to diversify cell fates, especially in nervous systems. In the Drosophila retina, stochastic expression of the PAS-bHLH transcription factor Spineless (Ss) controls photoreceptor subtype choice. In one randomly distributed subset of R7 photoreceptors, Ss activates Rhodopsin4 (Rh4) and represses Rhodopsin3 (Rh3); counterparts lacking Ss express Rh3 and repress Rh4. In the dorsal third region of the retina, the Iroquois Complex transcription factors induce Rh3 in Rh4-expressing R7s. Here, we show that Ss levels are controlled in a binary on/off manner throughout the retina yet are attenuated in the dorsal third region to allow Rh3 coexpression with Rh4. Whereas the sensitivity of rh3 repression to differences in Ss levels generates stochastic and regionalized patterns, the robustness of rh4 activation ensures its stochastic expression throughout the retina. Our findings show how stochastic and regional inputs are integrated to control photoreceptor subtype specification in the Drosophila retina.

Signaling pathways are often re-used during development in surprisingly different ways. The Hippo tumor suppressor pathway is best understood for its role in the control of growth. The pathway is also used in a very different context, in the Drosophila eye for the robust specification of R8 photoreceptor neuron subtypes, which complete their terminal differentiation by expressing light-sensing Rhodopsin (Rh) proteins. A double negative feedback loop between the Warts kinase of the Hippo pathway and the PH-domain growth regulator Melted regulates the choice between 'pale' R8 (pR8) fate defined by Rh5 expression and 'yellow' R8 (yR8) fate characterized by Rh6 expression. Here, we show that the gene encoding the homolog of human Nuclear respiratory factor 1, erect wing (ewg), is autonomously required to inhibit warts expression and to promote melted expression to specify pR8 subtype fate and induce Rh5. ewg mutants express Rh6 in most R8s due to ectopic warts expression. Further, ewg is continuously required to maintain repression of Rh6 in pR8s in aging flies. Our work shows that Ewg is a critical factor for the stable down-regulation of Hippo pathway activity to determine neuronal subtype fates. Neural-enriched factors, such as Ewg, may generally contribute to the contextual re-use of signaling pathways in post-mitotic neurons.

Signaling pathways are reused for multiple purposes in plant and animal development. The Hippo pathway in mammals and Drosophila coordinates proliferation and apoptosis via the coactivator and oncoprotein YAP/Yorkie (Yki), which is homeostatically regulated through negative feedback. In the Drosophila eye, cross-repression between the Hippo pathway kinase LATS/Warts (Wts) and growth regulator Melted generates mutually exclusive photoreceptor subtypes. Here, we show that this all-or-nothing neuronal differentiation results from Hippo pathway positive feedback: Yki both represses its negative regulator, warts, and promotes its positive regulator, melted. This postmitotic Hippo network behavior relies on a tissue-restricted transcription factor network-including a conserved Otx/Orthodenticle-Nrl/Traffic Jam feedforward module-that allows Warts-Yki-Melted to operate as a bistable switch. Altering feedback architecture provides an efficient mechanism to co-opt conserved signaling networks for diverse purposes in development and evolution.

Neuroepithelial cell proliferation must be carefully balanced with the transition to neuroblast (neural stem cell) to control neurogenesis. Here, we show that loss of the Drosophila microRNA mir-8 (the homolog of vertebrate miR-200 family) results in both excess proliferation and ectopic neuroblast transition. Unexpectedly, mir-8 is expressed in a subpopulation of optic-lobe-associated cortex glia that extend processes that ensheath the neuroepithelium, suggesting that glia cells communicate with the neuroepithelium. We provide evidence that miR-8-positive glia express Spitz, a transforming growth factor alpha (TGF-alpha)-like ligand that triggers epidermal growth factor receptor (EGFR) activation to promote neuroepithelial proliferation and neuroblast formation. Further, our experiments suggest that miR-8 ensures both a correct glial architecture and the spatiotemporal control of Spitz protein synthesis via direct binding to Spitz 3' UTR. Together, these results establish glial-derived cues as key regulatory elements in the control of neuroepithelial cell proliferation and the neuroblast transition.

In visual and olfactory sensory systems with high discriminatory power, each sensory neuron typically expresses one, or very few, sensory receptor genes, excluding all others. Recent studies have provided insights into the mechanisms that generate and maintain sensory receptor expression patterns. Here, we review how this is achieved in the fly retina and compare it with the mechanisms controlling sensory receptor expression patterns in the mouse retina and in the mouse and fly olfactory systems.

In the Drosophila optic lobes, the medulla processes visual information coming from inner photoreceptors R7 and R8 and from lamina neurons. It contains approximately 40,000 neurons belonging to more than 70 different types. Here we describe how precise temporal patterning of neural progenitors generates these different neural types. Five transcription factors-Homothorax, Eyeless, Sloppy paired, Dichaete and Tailless-are sequentially expressed in a temporal cascade in each of the medulla neuroblasts as they age. Loss of Eyeless, Sloppy paired or Dichaete blocks further progression of the temporal sequence. We provide evidence that this temporal sequence in neuroblasts, together with Notch-dependent binary fate choice, controls the diversification of the neuronal progeny. Although a temporal sequence of transcription factors had been identified in Drosophila embryonic neuroblasts, our work illustrates the generality of this strategy, with different sequences of transcription factors being used in different contexts.

PURPOSE: Surgical resection of midface neoplasms and subsequent reconstruction have been shown to have significant negative effects on quality of life (QOL). The purpose of this pilot study was to assess individuals' health-related QOL after maxillectomy and reconstruction with a prosthetic obturator. MATERIALS AND METHODS: The QOL of 25 of 43 patients who underwent maxillectomy and prosthetic obturator reconstruction at the University of California-San Francisco was assessed using 3 questionnaires: University of Washington Quality of Life version 4 (UWQOL), Obturator Functioning Scale (OFS), and Mental Health Inventory (MHI). RESULTS: The response rate to the QOL questionnaires was 92% (23 of 25 patients). Time elapsed from maxillectomy and prosthetic obturator reconstruction to the QOL survey response ranged from 0.3 to 6.6 years (mean, 2.7 years; standard deviation [SD], 1.9 years). The post-treatment mean QOL scores were 77.3 (SD, 13.6) for UWQOL, 72.0 (SD, 12.6) for OFS, and 4.5 (SD, 0.9) for Mental Health Inventory. Individuals who received adjuvant radiation scored lower for speech and appearance (OFS, P = .05, P = .03, respectively) as well as for saliva and overall QOL (UWQOL, P = .02, P = .08, respectively). There was a strong correlation between QOL scores in OFS and UWQOL questionnaires (r = 0.78, P < .001). CONCLUSION: The results of this pilot study suggest that postoperative radiation therapy was the strongest variable affecting QOL in patients with maxillectomy and prosthetic obturator reconstruction. There is further need for a multicenter trial with a larger sample to identify how factors affecting QOL of patients after maxillectomy might influence the choice of reconstruction.

PURPOSE: To analyze the effect of B0 field drift on multivoxel MR spectroscopic imaging and to propose an approach for its correction. THEORY AND METHODS: It is shown, both theoretically and in a phantom, that for approximately 30 min acquisitions a linear B0 drift ( approximately 0.1 ppm/h) will cause localization errors that can reach several voxels (centimeters) in the slower varying phase encoding directions. An efficient and unbiased estimator is proposed for tracking the drift by interleaving short ( approximately T2*), nonlocalized acquisitions on the nonsuppressed water each pulse repetition time, as shown in 10 volunteers at 1.5 and 3 T. RESULTS: The drift is shown to be predominantly linear in both the phantom and volunteers at both fields. The localization errors are observed and quantified in both phantom and volunteers. The unbiased estimator is shown to reliably track the instantaneous frequency in vivo despite only using a small portion of the FID. CONCLUSION: Contrary to single-voxel MR spectroscopy, where it leads to line broadening, field drift can lead to localization errors in the longer chemical shift imaging experiments. Fortunately, this drift can be obtained at a negligible cost to sequence timing, and corrected for in post processing.

We aim to improve segmentation through the use of machine learning tools during region agglomeration. We propose an active learning approach for performing hierarchical agglomerative segmentation from superpixels. Our method combines multiple features at all scales of the agglomerative process, works for data with an arbitrary number of dimensions, and scales to very large datasets. We advocate the use of variation of information to measure segmentation accuracy, particularly in 3D electron microscopy (EM) images of neural tissue, and using this metric demonstrate an improvement over competing algorithms in EM and natural images.

Animal behaviour arises from computations in neuronal circuits, but our understanding of these computations has been frustrated by the lack of detailed synaptic connection maps, or connectomes. For example, despite intensive investigations over half a century, the neuronal implementation of local motion detection in the insect visual system remains elusive. Here we develop a semi-automated pipeline using electron microscopy to reconstruct a connectome, containing 379 neurons and 8,637 chemical synaptic contacts, within the Drosophila optic medulla. By matching reconstructed neurons to examples from light microscopy, we assigned neurons to cell types and assembled a connectome of the repeating module of the medulla. Within this module, we identified cell types constituting a motion detection circuit, and showed that the connections onto individual motion-sensitive neurons in this circuit were consistent with their direction selectivity. Our results identify cellular targets for future functional investigations, and demonstrate that connectomes can provide key insights into neuronal computations.