Xeroderma pigmentosum: overview of pharmacology and novel therapeutic strategies for neurological symptoms

Xeroderma pigmentosum (XP) encompasses a group of rare diseases characterized in most cases by malfunction of nucleotide excision repair (NER), which results in an increased sensitivity to UV radiation in affected individuals. Approximately 25–30% of XP patients present with neurological symptoms, such as sensorineural deafness, mental deterioration and ataxia. Although it is known that dysfunctional DNA repair is the primary pathogenesis in XP, growing evidence suggests that mitochondrial pathophysiology may also occur. This appears to be secondary to dysfunctional NER but may contribute to the neurodegenerative process in these patients. The available pharmacological treatments in XP mostly target the dermal manifestations of the disease. In the present review, we outline how current understanding of the pathophysiology of XP could be used to develop novel therapies to counteract the neurological symptoms. Moreover, the coexistence of cancer and neurodegeneration present in XP led us to focus on possible new avenues targeting mitochondrial pathophysiology. Linked Articles: This article is part of a themed section on Mitochondrial Pharmacology: Featured Mechanisms and Approaches for Therapy Translation. To view the other articles in this section visit http://onlinelibrary.wiley.com/doi/10.1111/bph.v176.22/issuetoc.</p

TSPO interacts with VDAC1 and triggers a ROS-mediated inhibition of mitochondrial quality control

<p>The 18-kDa TSPO (translocator protein) localizes on the outer mitochondrial membrane (OMM) and participates in cholesterol transport. Here, we report that TSPO inhibits mitochondrial autophagy downstream of the PINK1-PARK2 pathway, preventing essential ubiquitination of proteins. TSPO abolishes mitochondrial relocation of SQSTM1/p62 (sequestosome 1), and consequently that of the autophagic marker LC3 (microtubule-associated protein 1 light chain 3), thus leading to an accumulation of dysfunctional mitochondria, altering the appearance of the network. Independent of cholesterol regulation, the modulation of mitophagy by TSPO is instead dependent on VDAC1 (voltage-dependent anion channel 1), to which TSPO binds, reducing mitochondrial coupling and promoting an overproduction of reactive oxygen species (ROS) that counteracts PARK2-mediated ubiquitination of proteins. These data identify TSPO as a novel element in the regulation of mitochondrial quality control by autophagy, and demonstrate the importance for cell homeostasis of its expression ratio with VDAC1.</p

Whole-cell current in R29A transfected HEK cells.

<p>Family of whole-currents for WT (A) and R29A (B) transfected HEK cells (top panels). Voltage steps lasting 800 ms from holding potential of -30 mV to membrane potential of -80 to +100 mV (20 mV step increment). The middle panels depict the whole-cell currents after perfusion of 50 µM of IAA94. The bottom panels represent the IAA94-sensitive currents obtained by subtraction of the middle panel current from the upper panel current. Note that the IAA94-sensitive currents are plotted on a different scale with the scale bars on the right hand side of the figure. (C) an example of current/voltage relationship of the IAA94-sensitive current from a WT (□) and a R29A (○) transfected HEK cell. (D) Averaged G/V plots from IAA94-sensitive current of WT (□) and R29A mutant (○), from 5 independent experiments.</p

CLIC1 ion channel activity from Tip-Dip bilayer experiments.

<p>Current recordings from -80 to +80 mV, 20 mV interval, are shown in (A) for wild type CLIC1 (left), K37A (center) and R29A (right) CLIC1 protein. The upper panel of <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0074523#pone-0074523-g001" target="_blank">Figure 1B</a> depicts the single-channel current-voltage data for WT (□) and K37A (○) CLIC1 proteins. The average single-channel conductance differs between WT and K37A, calculated as 30.1 ± 0.2 and 42.4 ± 0.2 pS, respectively (n = 5, p < 0.001). In contrast, the channel open probability of the K37A mutation is very similar to the WT (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0074523#pone-0074523-g001" target="_blank">Figure 1B</a>, lower panel). The current/voltage relationships for both WT (□) and R29A CLIC1 (○) are shown in (C). The single-channel conductance (upper panel), is 30.1 ± 0.2 and 29 ± 0.2 pS for WT and R29A, respectively. The average open probability for WT (□) and R29A mutated protein (○) is shown in the lower panel.</p

Endogenous CLIC1 is not expressed on the plasma membrane of untransfected HEK cells.

<p>(A) Family of currents for an untransfected HEK293 cell at varying applied voltages from -60 to +60 mV with 20 mV increment. Top panel: whole cell current in resting conditions; middle panel: after IAA94 perfusion; lower panel: IAA94-sensitive (CLIC1-mediated) current obtained from subtraction. (B) i/V curve of an IAA94-sensitive current in WT CLIC1 transfected HEK293 cell (□), and in an untransfected HEK293 cell (○). (C) Plot of the average of the IAA94-sensitive current as a percentage of the control current of untransfected HEK cells (n=5). Data are shown as mean ± SEM. CLIC1-mediated current is completely absent in untransfected HEK cells.</p

Open (τ_open) and close (τ_close) time constants of the WT and K37A CLIC1 ion channel.

<p>In panels (A) and (B), membrane potentials were held at +35 mV while panels (C) and (D) concern cell membrane potentials held at -5 mV for WT (A, C) and K37A (B, D) CLIC1 transfected HEK cells. (A) to (D) shows the open (left) and close (right) time distributions for each condition. Four seconds of single channel recordings appear as inserts in the corresponding panels for each condition. The open and close time distribution histograms were fitted by a double exponential decay function and plotted on a semi-logarithmic scale. Panels (E) and (F) depict open and close time distributions as a function of membrane potential for WT (□) and K37A (○) transfected HEK cells.</p

Cell-attached recordings of HEK cells transiently transfected with WT CLIC1 and R29A CLIC1 protein.

<p>Single-channel current traces are shown for WT (A) and R29A (B) transfected cells. The voltage steps are from -5 to + 25 mV (10 mV step increment). Single-channel current/voltage plots are shown in (C) for WT (□) and R29A (○). Average single-channel conductance of 12.3 ± 0.1 pS for WT and 13.1 ± 0.3 pS for R29A and an extrapolated reversal potential of -63 ± 0.4 mV and -61 ± 0.6 mV, respectively. The open probability obtained at each membrane potential is shown in (D) for WT (□) and R29A (○).</p

Cell-attached recordings of HEK cells transiently transfected with WT and K37A CLIC1 protein.

<p>Single-channel current traces are shown for WT (A) and K37A (B) CLIC1 transfected cells. The membrane voltage was clamped at different values (indicated on the left of each trace) in 10 mV step increments. Single-channel current/voltage plots are shown in (C) for WT (□) and K37A (○). Average single-channel conductance of 12.1 ± 0.6 pS for WT and 17.4 ± 0.8 pS for K37A with an extrapolated reversal potential of -58 ± 0.7 mV and -56 ± 1.2 mV, respectively, was calculated. The open probability obtained at each membrane potential is shown in (D) for WT (□) and K37A (○).</p

Additional file 3: Figure S3. of Glial cells are functionally impaired in juvenile neuronal ceroid lipofuscinosis and detrimental to neurons

Composition of Microglial Cultures. Primary cortical microglial cultures generated from P2–4 wild type (WT) and Cln3-deficient (Cln3 −/− ) mice were immunostained with CD68 to identify microglia, O4 to identify oligodendrocytes, TuJ1 to identify neurons and GFAP to identify astrocytes. DAPI was used to visualize all nuclei. Practically all cells were CD68 expressing microglial cells (A), with virtually no cells expressing GFAP or O4 (B). Scale bar = 20 μm. (TIFF 8572 kb

Additional file 6: Figure S5. of Glial cells are functionally impaired in juvenile neuronal ceroid lipofuscinosis and detrimental to neurons

An intact actin cytoskeleton is essential for glutathione secretion. To study the importance of the actin cytoskeleton for GSH secretion in astrocytes Cytochalasin D (1uM) was added to wild type (WT) astrocytes for 30 min prior to the start of the 8 h period over which the accumulation of secreted GSH in the medium was measured. Cells were then fixed and the actin cytoskeleton visualized with phalloidin. DAPI was used to visualize all nuclei. (A) Cytochalasin D clearly disrupted the F-actin filament organization in WT astrocytes. (B) Perturbing actin cytoskeletal polymerization significantly inhibited GSH secretion by WT astrocytes. Scale bar in (A) = 10 um. (TIFF 13687 kb