Research interests

Genetic Analysis of Cerebellar Development and Function

The focus of our laboratory is the elucidation of the molecular mechanisms underlying the development and maintenance of neurons in the brain. As described below, we have taken a forward genetic approach to identify novel pathways involved in these processes.

Netrin signaling in CNS development

Mice homozygous for mutations in the rostral cerebellar malformation (rcm) gene (now identified as Unc5c) exhibit cerebellar and midbrain defects. The cerebellum of mutants is reduced in both size and number of folia, and the midbrain and brainstem contain ectopic cerebellar neurons. We found that the rcm cDNA encodes a transmembrane receptor of the immunoglobulin superfamily that is highly similar to the Caenorhabditis elegans protein UNC-5, which is essential for dorsal guidance of pioneer axons. UNC-5 is also necessary for movement of cells away from the netrin ligand, which is encoded by the unc-6 gene. Our studies have shown that this netrin receptor is necessary for recognition of the anterior border of the cerebellum by migrating granule cell precursors during embryogenesis, and for recognition of the ventral boundary of the inner granule cell layer in the lateral regions of the cerebellum by radially migrating postnatal granule cells. In addition to neuronal migration abnormalities, we have found corticospinal tract defects in both Unc5crcm/Unc5crcm mice and mice homozygous for a mutation in the netrin receptor, deleted in colorectal carcinoma, demonstrating a role for these receptors in navigations of these axons.

Recently, we have observed that Unc5c mutants on an inbred genetic background die shortly after birth, apparently due to respiratory problems. In contrast, mutant mice on a segregating background live a normal life span. Analysis of mutant embryos on this inbred background revealed that Unc5c is necessary for guidance of the phrenic nerve, which normally innervates the diaphragm muscle, and trochlear motor axons defining a novel role for mammalian netrins and their receptors in motor axon guidance. We are currently analyzing F2 mice in an effort to genetically define the modifier genes that influence guidance of these axons. 

Oxidative stress, cell cycle and neurodegeneration

We have used a forward genetic approach to understanding the molecular mechanisms underlying neuron loss in the aging mammalian brain. To identify the molecular mechanisms for granule cell survival, we are studying the spontaneous mutant, harlequin (Hq). Adult Hq mutant mice develop progressive ataxia and retinal degeneration. Cell loss in the cerebellum and retina peaks at 5-7 months. Large numbers of mature retinal neurons and cerebellar granule cells, many of which are undergoing apoptosis, are in the S phase of the cell cycle in these mice.

Using a positional cloning approach, we have identified the Hq mutation as a proviral insertion in the apoptosis-inducing factor (Aif) gene (formally designated programmed cell death 8, Pdcd8), causing an approximate 80 percent reduction in AIF expression. AIF is a mitochondrial oxidoreductase that contains a pyridine nucleotide-disulphide oxidoreductase subunit similar to regulators of plant and bacterial hydrogen peroxide scavengers. Although the function of AIF within the mitochondria is unknown, AIF has been previously shown to cause apoptosis when it translocates to the nucleus.

Our studies show that reduced levels of AIF lead to oxidative stress in the retina and cerebellum. Further, we demonstrate that it is these stressed neurons that reenter the cell cycle. Thus, these data demonstrate that oxidative stress can mediate neuronal cell cycle reentry and subsequent neuronal apoptosis in the adult central nervous system (CNS). We are currently evaluating signaling pathways that are regulated by oxidative stress and may result in this aberrant reentry into the cell cycle and subsequent apoptosis. Lastly, we are also investigating a modifier gene that we have identified that suppresses neurodegeneration in both Hq mutant neurons, and in neurons in a new spontaneous mutant that has oxidative stress-induced neuron death. The identification of this gene will give additional insights into the means by which oxidative stress induces apoptosis and cell cycle re-entry.

Protein misfolding and neurodegeneration

Oligomerization and the formation of aggregates of misfolded proteins are common to many genetic and sporadic forms of neurodegenerative diseases. Some of these misfolded proteins are due to mutations directly within disease-related proteins, such as in the polyglutamine expansion diseases and some forms of familial Alzheimer’s disease. However, the mechanisms underlying protein misfolding in many neurodegenerative diseases remain unknown. We have identified two loss of function models of Purkinje cell degeneration associated with the accumulation of misfolded proteins. Mice homozygous for the woozy (wz) mutation develop ataxia between 3 and 4 months of age concomitant with Purkinje cell loss. Prior to degeneration, ubiquitinated protein accumulations are found in the endoplasmic reticulum (ER) and nucleus in these neurons. These abnormal protein accumulations induce the cellular response known as the unfolded protein response, which acts to help restore ER homeostasis. By positional cloning, we identified the wz mutation in the Sil1 gene, which encodes a co-chaperone of the ER chaperone and ER-stress transducer, BiP. These data suggest that the adenine nucleotide exchange function of SIL1 is necessary for the chaperone function of BiP, but not for ER-stress function of BiP. Recently SIL1 mutations were found in several families with Marinesco-Sjögren syndrome, which is associated with cerebellar ataxia. Thus the woozy mutant mouse will be an excellent model for this syndrome.

Purkinje cell loss in mice homozygous for the spontaneous sticky (sti) mutation is associated with accumulation of ubiquitinated proteins in the cytoplasm, ER, and nucleolus. We established that the sti molecular defect is a point mutation in the editing domain of alanyl tRNA synthetase (AARS). The aminoacyl tRNA synthetases establish the genetic code where each amino acid is linked to its cognate tRNA that bears the anticodon triplet of the code. The high accuracy of protein translation is largely due to the precision of these aminoacylation reactions, and much of this accuracy resides in the editing domains of these synthetases which can clear misactivated amino acids or mischarged tRNAs.

In collaboration with Paul Schimmel’s laboratory (The Scripps Research Institute), we have demonstrated that the sticky mutation causes an increase in mischarged tRNAAla. This in turn leads to random misincorporation of amino acids at Ala codons, ultimately leading to the production of unfolded, heterogenous proteins. The loss of translational fidelity in sti mutant mice provides an exciting new mechanism underlying neurodegeneration.

Recently we have identified a dominant modifier gene called Msti that suppresses neurodegeneration in sti mutant mice. Currently we are performing experiments to delineate the protective role of Msti in sti/sti Purkinje cells. We are also testing whether Msti protects against other misfolded protein-associated neurodegenerative disorders. 

Publications

• Ackerman SL, Cox GA. 2008. From ER to Eph receptors: new roles for VAP fragments. Cell 133:949-951. (Preview)
• Hu Z, Shanker S, Maclean JA 2nd, Ackerman SL, Wilkinson MF. 2008. The RHOX5 homeodomain protein mediates transcriptional repression of the netrin-1 receptor gene Unc5c. J Biol Chem. 283:3866-3876.
• Ishimura R, Martin GR, Ackerman SL. 2008. Loss of apoptosis inducing factor results in cell type-specific neurogenesis defects. J Neurosci. 28:4938-4948.
• Renaud J, Kerjan G, Sumita I, Zagar Y, Georget V, Kim D, Fouquet C, Suda K, Sanbo M, Suto F, Ackerman SL, Mitchell KJ, Fujisawa H, Chédotal A. 2008. Plexin-A2 and its ligand Sema6A control nucleus-centosome coupling in migrating cerebellar granule cells. Nat Neurosci. 11:440-449.
• Bernet A, Mazelin L, Coissieux MM, Gadot N, Ackerman SL, Scoazec JY, Mehlen P. 2007. Inactivation of the UNC5C Netrin-1 receptor is associated with tumor progression in colorectal malignancies. Gastroenterology 133:1840-1848.
• Dillon AK, Jevince AR, Hinck L, Ackerman SL, Lu X, Tessier-Lavigne M, Kaprielian Z. 2007. UNC5C is required for spinal accessory motor neuron development. Mol Cell Neurosci. 35:482-489.
• Xie Y, Hong Y, Ma X-Y, Ren XR, Ackerman S, Mei L, Xiong W-C. 2006. DCC-dependent phospholipase C signaling in netrin-1 induced neurite elongation. J Biol Chem 281:2605-2611.
• Watanabe K, Tamamaki N, Furuta T, Ackerman SL, Ikenaka K, Ono K. 2006. Dorsally derived netrin-1 provides an inhibitory cue and elaborates the 'waiting period' for primary sensory axons in the developing spinal cord. Development 133:1379-1387.
• van Empel VPM, Bertrand AT, van Oort RJ, van der Nagel R, Engelen M, van Rijen HV, Doevendans PA, Crijns HJ, Ackerman SL, Sluiter W, De Windt LJ. 2006. EUK-8, a superoxide dismutase and catalase mimetic, reduces cardiac oxidative stress and ameliorates pressure overload-induced heart failure in the harlequin mouse mutant. J Am Coll Cardiol 48:824-832.
• Lee JW, Beebe K, Nangle LA, Jang J, Longo-Guess CM, Cook SA, Davisson MT, Schimmel, Ackerman SL. 2006. Editing-defective tRNA synthetase causes protein misfolding and neurodegeneration. Nature 443:50-55.
• Burgess RW, Jucius TJ, Ackerman SL. 2006. Motor axon guidance of the mammalian trochlear and phrenic nerves: Dependence on the netrin receptor Unc5c and modifier loci. J Neurosci 26:4455-4459.
• Bock NA, Kovacevic N, Lipina TV, Roder JC, Ackerman SL, Henkelman RM. 2006. In vivo magnetic resonance imaging and semiautomated image analysis extend the brain phenotype for cdf/cdf mice. J Neurosci 26:4455-4459.
• Zhao L, Ackerman SL. 2006. ER stress in disease. Curr Opin Cel Biol 18:1-9.
• van Empel VPM, Bertrand AT, van der Nagel R, Kostin S, Doevendans PA, Crijns HJ, de Wit E, Sluiter W, Ackerman SL, De Windt LJ. 2005. Downregulation of apoptosis-inducing factor in harlequin mutant mice sensitizes the myocardium to oxidative stress-related cell death and pressure overload-induced decompensation. Circ Res 96: 92-101.
• Xie Y, Ding Y-Q, Hong Y, Feng Z, Navarre S, Xi C-X, Zhu X-J, Wang C-L, Ackerman SL, Kozlowski D, Mei L, Xiong W-C. 2005. Phosphatidylinositol transfer protein-α in netrin-1-induced PLC signalling and neurite outgrowth. Nat Cell Biol 7:1124-1132.
• Zhao L, Longo-Guess C, Harris BS, Lee JW, Ackerman SL. 2005. Protein accumulation and neurodegeneration in the woozy mutant mouse is caused by disruption of SIL1, a cochaperone of BiP. Nat Genet 37:974-979.