Eye Research Institute
Eye Research Institute Website
Mitton Lab File Sharing (password req'd.)
Science of Vision - Bio 487 Student Course Page, Mitton Lectures
M.O.P.'s
Link for Peer-Reviewed BioMedical Articles:
(Mothers
of pre-schoolers & Home Educators)
|
|
| Our Latest Research
Publication Simpanya MF, Wistow G, David LL, Giblin FJ, Mitton KP*. (2008) Expressed sequence tag analysis of guinea pig (Cavia porcellus) eye tissues for NEIBank. Mol Vision2008; 14:2413-2427 Open Access HERE - This paper documents the first extensive EST analysis of isolated guinea pig eye tissues, and their inclusion in the NEIBank data base. EST sequences from this work represent the first large submission of ESTs and EST derived cDNAs into GenBank for the guinea pig. This data permitted us to construct general gene structures for many key vision genes of the retina and lens, for comparison to other mammals, including humans. This large set of ESTs (cDNA sequences) are now available to confirm intron/exon structures in the recently sequenced guinea-pig genome. Mali
RS, Peng G-H, Zhang X, Dang L, Chen S, Mitton KP*. (2008) FIZ1 is part
of the regulatory protein complex on active photoreceptor-specific gene
promoters in vivo.
BMC Molecular Biology 9, 87 Open Access HERE - This paper demonstrates that FIZ1 protein is recruited to several photoreceptor gene promoter complexes as the genes become active, using ChIP analysis. Other novel findings are the capacity for direct protein interaction between FIZ1 and CRX. We also used Chromatin immunoprecipitation (ChIP) to detect actively transcribing RNA-Polymerase-II moving within the Rhodopsin gene's transcriptional region , within neural retina tissue. Mali RS, Zhang X, Hoerauf W, Doyle S, Devitt J, Loffreda-Wren J, and Mitton KP*. (2007) FIZ1 is Expressed During Photoreceptor Maturation, and Synergizes with NRL and CRX at Rod-Specific Promoters, in vitro. Exp Eye Res 84, 349-360 - This paper shows that the expression of FIZ1 protein increases as the neural retina matures. FIZ1 appears in higher concentrations just when key photoreceptor genes are expressed, like Rhodopsin. Furthermore, FIZ1 demonstrates the ability to synergize with NRL to increase NRL's activation of two important rod-specific gene promoters; the Rhodopsin gene promoter and the PDE6B gene promoter. |
| Other Recent Research Collaborations Discovery of the function of the human NR2E3 gene product (enhanced S-cone syndrome). NR2E3, or photoreceptor specific nuclear receptor (PNR), was already known to be mutated in human retinal disease (enhanced S-cone syndrome), but its function was not known. Dr. Mitton used yeast two-hybrid interaction trapping to discover the interation of NR1D1 with NR2E3 while completing post-doctoral training in Anand Swaroop's laboratory at the University of Michigan. We continued to collaborate with Dr Swaroop to test this interaction using yeast two-hybrid technology (here at Oakland University), and to assist with the design of other experiments. Anand Swaroop is now at the National Eye Insitute - NIH. This paper documents that NR2E3 is only
expressed in rod-photoreceptors (not cones), and that both NR2E3 and NR1D1
interact with rod-specific genes (ie. rhodopsin). Our discovery of NR1D1 association
with NR2E3 is the first clue of how the cell's biological clock can regulate
the expression of rod-specific genes over a 24 hour cycle. In humans, rhodopsin
gene expression levels are highest at mid-day and lowest at midnight. Examinations
of transcription factors known to control rhodopsin gene expression
failed to explain this variation. However, NR1D1 is well known as an important
member of the cell's biological clock; NR1D1 integrates the positive and negative
feedback loops of an intracellular genetic clock in every cell. NR1D1 provides
a physical link between rod-specific gene expression and cell' s biological
clock, our first clue to this puzzle.Cheng et
al., in this issue. Support Oakland's Vision Science Research and Training of Future Scientists.. You may not be a scientist, but you can support and contribute to vision science research and training at Oakland University. Since 2002, the Mitton lab has directly provided advanced research training to over 15 Oakland University undergraduate students. Not practice lab work, real research! Four of them are co-authors on our 2007 EER paper. Some of these students are now in graduate Ph.D. programs in Human Genetics and Neuroscience, as well as Medical and Optometry School. Resources are the limiting factor for most biomedical research programs. Most of us have ideas we just have to keep on the shelf. There are not enough funds for the hands, instruments and biochemicals to work on them all. If you are curious about supporting such endeavours, right here in Oakland County, you can come visit the ERI and we will be happy to show you our institute. If you would like to support our research and training programs, contact Dr. Mitton at mitton@oakland.edu |
Assistant Professor of Biomedical Sciences
Eye Research Institute
Oakland University
Control of Gene Expression Laboratory
Systems Biology with a Focus
Rochester MI 48309
mitton@oakland.edu
| Why Our NIH-funded
Research Is Important. Rhodopsin is the light detecting protein of our eyes. It is only made in rod-photoreceptors because the rhodopsin gene is only expressed in rods. Many retinal diseases in humans are due to mutations in genes like the Rhodopsin gene. Diseases such as Retinitis Pigmentosa (RP). RP can also result when the Rhodopsin gene is normal, but mutations alter the genes that code for special proteins (called transcription factors) that switch and control the Rhodopsin gene. Like the saying goes, "Too much or too little of a good thing can hurt you." Having too much, or too little, normal Rhodopsin can be as harmful to your photoreceptor cells as having a disfunctional Rhodopsin protein. Transcription factors act as switches to turn genes on
or off (called gene expression). My lab has discovered key interactions
between these protein-switches that help us understand how the Rhodopsin
gene is expressed in rod photoreceptor cells. Currently the focus of my lab is a novel protein called FIZ1. I discovered this protein interacting with a rod photoreceptor-specific transcription factor called NRL. NRL is important for turning on genes that are required for retinal precursor cells to become working photoreceptors during development of the retina. Mutations in the NRL gene also cause Retinitis Pigmentosa. Dr. Anand Swaroop's lab, at the University of Michigan, has shown that NRL is an essential protein to have normal rod-cells in the retina. In my lab, we currently believe that FIZ1 is part of an interface between cell-specific transcription factors, like NRL, and more ubiquitous proteins that support the precise expression level of genes activated by NRL (in the case of rod-photorecptors). We MUST fully understand the regulatory networks that result in the formation of normal photoreceptor cells to support the design of thereapies that are SAFE. Gene therapies can, and already have, inadvertently introduce new problems with the good intention of correcting the original defect. Ignorance of the full system of interactions only increases the chance of such unfortunate results. Information that we discover, combined with that of many other laboratories around the world, will improve our ability to understand the molecular reasons for a disease and help design safe therapies.How do we map these regulatory protein networks? There are tens of thousands of genes and proteins expressed in any one cell. How can you map all the interactions that lead to one place, like the control of Rhodopsin expression? We use a Systems Biology approach, but with a focus. In other words we use some powerful information methods to get ideas about gene and protein interactions, using bioinfomatics tools. We also use direct molecular biology methods to trace and discover new protein interactions (the "focus). The later methods include techniques like the yeast-2-hybrid protein interaction trap. That method permits us to take one important protein, and survey over a million retinal cDNA-library clones for unknown proteins that bind with our protein of interest. I found several important protein interactions this way, which involve transcription factors involved in retinal gene control and retinal disease. Most recently, we have completed the first ChIP-on-Chip analysis to map the association of RNA-Polymerase-II at 26,000 genes during maturation of the neural retina. This provides a novel way to directly detect changes in gene activation state in neural tissues in vivo. I continue to explore, with FIZ1 as a network focus, and teach my research skills to undergraduate students, post-docs, and ophthalmology residents at Oakland University and the William Beaumont Hospital. SEEING RHODOPSIN.... 
This picture shows immunofluorescent
detection of the rhodopsin protein in the mouse retina (green). Chromosoms
(DNA) in the nuclei of retinal cells are labeled with a specific stain (blue).
Each color, (blue, green) was captured in a separate digital image with a
digital light fluorescent microscroscopy system. The color layers were merged
for this final image. Rhodopsin (green) is located in long specialized structures
of the photoreceptor cells called the outer segments. The large layer of
blue above the green is the nuclei of the photoreceptors (called the outer
nuclear layer). (by Widmann Hoerauf "Woody" in the Mitton Lab 2004/5) |