Warren G. Tourtellotte

Bax-Deficient Mice with Lymphoid Hyperplasia and Male Germ Cell Death

BAX, a heterodimeric partner of BCL2, counters BCL2 and promotes apoptosis in gain-of-function experiments. A Bax knockout mouse was generated that proved viable but displayed lineage-specific aberrations in cell death. Thymocytes and B cells in this mouse displayed hyperplasia, and Bax-deficient ovaries contained unusual atretic follicles with excess granulosa cells. In contrast, Bax-deficient males were infertile as a result of disordered seminiferous tubules with an accumulation of atypical premeiotic germ cells, but no mature haploid sperm. Multinucleated giant cells and dysplastic cells accompanied massive cell death. Thus, the loss of Bax results in hyperplasia or hypoplasia, depending on the cellular context.

The EGR family of transcription-regulatory factors: progress at the interface of molecular and systems neuroscience

The EGR family of transcription regulatory factors, which is implicated in orchestrating the changes in gene expression that underlie neuronal plasticity, has attracted the attention of both molecular and systems neuroscientists. In this article, the advances made in both these fields of research are reviewed. Recent systems-based studies underscore the remarkable sensitivity and specificity of the induction of the expression of genes encoding EGR-family members in naturally occurring plasticity paradigms. However, they also challenge conventional views of the role of this family in plasticity. Recent molecular studies have identified the gonadotropin subunit, luteinizing hormone beta, as an EGR1-regulated gene in vivo and uncovered an essential role for EGR3 in muscle-spindle development. In addition, the discovery of novel proteins that are capable of suppressing EGR-mediated transcription cast doubt over the prevalent assumption that changes in EGR mRNA or protein levels provide an accurate measure of EGR-driven transcriptional activity.

Sensory ataxia and muscle spindle agenesis in mice lacking the transcription factor Egr3

Muscle spindles are skeletal muscle sensory organs that provide axial and limb position information (proprioception) to the central nervous system. Spindles consist of encapsulated muscle fibers (intrafusal fibers) that are innervated by specialized motor and sensory axons. Although the molecular mechanisms involved in spindle ontogeny are poorly understood, the innervation of a subset of developing myotubes (type I) by peripheral sensory afferents (group Ia) is a critical event for inducing intrafusal fiber differentiation and subsequent spindle formation. The Egr family of zinc-finger transcription factors, whose members include Egr1 (NGFI-A), Egr2 (Krox-20), Egr3 and Egr4 (NGFI-C), are thought to regulate critical genetic programs involved in cellular growth and differentiation (refs 4, 5, 6, 7, 8 and W.G.T. et al., manuscript submitted). Mice deficient in Egr3 were generated by gene targeting and had gait ataxia, increased frequency of perinatal mortality, scoliosis, resting tremors and ptosis. Although extrafusal skeletal muscle fibers appeared normal, Egr3-deficient animals lacked muscle spindles, a finding that is consistent with their profound gait ataxia. Egr3 was highly expressed in developing muscle spindles, but not in Ia afferent neurons or their terminals during developmental periods that coincided with the induction of spindle morphogenesis by sensory afferent axons. These results indicate that type I myotubes are dependent upon Egr3-mediated transcription for proper spindle development.

The Neuroplasticity-Associated Arc Gene Is a Direct Transcriptional Target of Early Growth Response (Egr) Transcription Factors

ABSTRACT Early growth response (Egr) transcription factors (Egr1 to Egr4) are synaptic activity-inducible immediate early genes (IEGs) that regulate some aspects of synaptic plasticity-related to learning and memory, yet the target genes regulated by them are unknown. In particular, Egr1 is essential for persistence of late-phase long-term potentiation (L-LTP), for hippocampus-dependent long-term memory formation, and for reconsolidation of previously established memories. Here, we show that Egr1 and Egr3 directly regulate the plasticity-associated activity-regulated cytoskeletal-related (Arc) gene, a synaptic activity-induced effector molecule which is also required for L-LTP and hippocampus-dependent learning and memory processing. Moreover, Egr1-deficient and Egr3-deficient mice lack Arc protein in a subpopulation of neurons, while mice lacking both Egr1 and Egr3 lack Arc in all neurons. Thus, Egr1 and Egr3 can indirectly modulate synaptic plasticity by directly regulating Arc and the plasticity mechanisms it mediates in recently activated synapses.

ALS-associated mutation FUS-R521C causes DNA damage and RNA splicing defects

Autosomal dominant mutations of the RNA/DNA binding protein FUS are linked to familial amyotrophic lateral sclerosis (FALS); however, it is not clear how FUS mutations cause neurodegeneration. Using transgenic mice expressing a common FALS-associated FUS mutation (FUS-R521C mice), we found that mutant FUS proteins formed a stable complex with WT FUS proteins and interfered with the normal interactions between FUS and histone deacetylase 1 (HDAC1). Consequently, FUS-R521C mice exhibited evidence of DNA damage as well as profound dendritic and synaptic phenotypes in brain and spinal cord. To provide insights into these defects, we screened neural genes for nucleotide oxidation and identified brain-derived neurotrophic factor (Bdnf) as a target of FUS-R521C-associated DNA damage and RNA splicing defects in mice. Compared with WT FUS, mutant FUS-R521C proteins formed a more stable complex with Bdnf RNA in electrophoretic mobility shift assays. Stabilization of the FUS/Bdnf RNA complex contributed to Bdnf splicing defects and impaired BDNF signaling through receptor TrkB. Exogenous BDNF only partially restored dendrite phenotype in FUS-R521C neurons, suggesting that BDNF-independent mechanisms may contribute to the defects in these neurons. Indeed, RNA-seq analyses of FUS-R521C spinal cords revealed additional transcription and splicing defects in genes that regulate dendritic growth and synaptic functions. Together, our results provide insight into how gain-of-function FUS mutations affect critical neuronal functions.

Fibrosis in systemic sclerosis: Emerging concepts and implications for targeted therapy

Systemic sclerosis (SSc) is a complex and incompletely understood disease associated with fibrosis in multiple organs. Recent findings identify transforming growth factor-ß (TGF-ß), Wnt ligands, toll-like receptor-mediated signaling, hypoxia, type I interferon, type 2 immune responses and mechanical stress as extracellular cues that modulate fibroblast function and differentiation, and as potential targets for therapy. Moreover, fibrillin-1 has a major role in storing and regulating the bioavailability of TGF-ß and other cytokines, and fibrillin-1 mutations are implicated in a congenital form of scleroderma called stiff skin syndrome. Fibrosis is due not only to the activation of tissue-resident fibroblasts and their transdifferentiation into myofibroblasts, but also the differentiation of bone marrow-derived fibrocytes, and transition of endothelial and epithelial cells, pericytes and adipocytes into activated mesenchymal cells. These responses are modulated by signaling mediators and microRNAs that amplify or inhibit TGF-ß and Wnt signaling. Gain-of-function and loss-of-function abnormalities of these mediators may account for the characteristic activated phenotype of SSc fibroblasts. The nuclear orphan receptor PPAR-γ plays a particularly important role in limiting the duration and intensity of fibroblast activation and differentiation, and impaired PPAR-γ expression or function in SSc may underlie the uncontrolled progression of fibrosis. Identifying the perturbations in signaling pathways, mediators and differentiation programs that are responsible for SSc tissue damage allows their selective targeting. This in turn opens the door for therapies utilizing novel compounds, or drug repurposing by innovative uses of already-approved drugs. In view of the heterogeneous clinical presentation and unpredictable course of SSc, as well as its complex pathogenesis, only robust clinical trials incorporating the judicious application of biomarkers will be able to clarify the clinical utility of these innovative approaches.

microRNA-21 Regulates Astrocytic Response Following Spinal Cord Injury

Astrogliosis following spinal cord injury (SCI) involves an early hypertrophic response that serves to repair damaged blood–brain barrier and a subsequent hyperplastic response that results in a dense scar that impedes axon regeneration. The mechanisms regulating these two phases of astrogliosis are beginning to be elucidated. In this study, we found that microRNA-21 (miR-21) increases in a time-dependent manner following SCI in mouse. Astrocytes adjacent to the lesion area express high levels of miR-21 whereas astrocytes in uninjured spinal cord express low levels of miR-21. To study the role of miR-21 in astrocytes after SCI, transgenic mice were generated that conditionally overexpress either the primary miR-21 transcript in astrocytes or a miRNA sponge designed to inhibit miR-21 function. Overexpression of miR-21 in astrocytes attenuated the hypertrophic response to SCI. Conversely, expression of the miR-21 sponge augmented the hypertrophic phenotype, even in chronic stages of SCI recovery when astrocytes have normally become smaller in size with fine processes. Inhibition of miR-21 function in astrocytes also resulted in increased axon density within the lesion site. These findings demonstrate a novel role for miR-21 in regulating astrocytic hypertrophy and glial scar progression after SCI, and suggest miR-21 as a potential therapeutic target for manipulating gliosis and enhancing functional outcome.

Degradation of mouse locomotor pattern in the absence of proprioceptive sensory feedback

Significance Terrestrial locomotion is thought to be generated by the actions of a circuit of interconnected interneurons (central pattern generator) in the spinal cord that drive the patterned activity of pools of motor neurons, causing sequential contraction of dozens of leg muscles. Sensory feedback exerts a strong modulatory influence on this pattern; nevertheless, it remains unclear whether sensory feedback also plays a role in the generation of the normal locomotor pattern. Through the use of a combination of electrophysiology, behavior, and mouse genetics, we provide evidence that the absence of proprioceptive sensory feedback degrades locomotor pattern, indicating that proprioceptive feedback is required for the construction of locomotor pattern. Mammalian locomotor programs are thought to be directed by the actions of spinal interneuron circuits collectively referred to as “central pattern generators.” The contribution of proprioceptive sensory feedback to the coordination of locomotor activity remains less clear. We have analyzed changes in mouse locomotor pattern under conditions in which proprioceptive feedback is attenuated genetically and biomechanically. We find that locomotor pattern degrades upon elimination of proprioceptive feedback from muscle spindles and Golgi tendon organs. The degradation of locomotor pattern is manifest as the loss of interjoint coordination and alternation of flexor and extensor muscles. Group Ia/II sensory feedback from muscle spindles has a predominant influence in patterning the activity of flexor muscles, whereas the redundant activities of group Ia/II and group Ib afferents appear to determine the pattern of extensor muscle firing. These findings establish a role for proprioceptive feedback in the control of fundamental aspects of mammalian locomotor behavior.

Muscle spindle-derived neurotrophin 3 regulates synaptic connectivity between muscle sensory and motor neurons

Ia afferents induce the formation of muscle spindles prenatally and maintain them postnatally. To address whether spindles, in turn, regulate the function of Ia afferents, we examined Egr3-null mutant mice (Egr3−/−), in which muscle spindles degenerate progressively after birth. Egr3−/− mice develop gait ataxia, scoliosis, resting tremors, and ptosis, suggesting a defect in proprioception. Despite the normal morphological appearance of peripheral and central sensory projections, we observed a profound functional deficit in the strength of sensory–motor connections in Egr3−/− mice. Muscle spindles in Egr3−/− mice do not express NT3. Intramuscular injections of NT3 to Egr3−/− mice during the postnatal period restored sensory–motor connections. Thus, NT3 derived from muscle spindles regulates the synaptic connectivity between muscle sensory and motor neurons.

Myofibroblasts in Murine Cutaneous Fibrosis Originate From Adiponectin‐Positive Intradermal Progenitors

Accumulation of myofibroblasts in fibrotic skin is a hallmark of systemic sclerosis (SSc; scleroderma), but the origins of these cells remain unknown. Because loss of intradermal adipose tissue is a consistent feature of cutaneous fibrosis, we sought to examine the hypothesis that myofibroblasts populating fibrotic dermis derive from adipocytic progenitors.