Matthew D. Benson

Palmitoylated TMX and calnexin target to the mitochondria-associated membrane

The mitochondria‐associated membrane (MAM) is a domain of the endoplasmic reticulum (ER) that mediates the exchange of ions, lipids and metabolites between the ER and mitochondria. ER chaperones and oxidoreductases are critical components of the MAM. However, the localization motifs and mechanisms for most MAM proteins have remained elusive. Using two highly related ER oxidoreductases as a model system, we now show that palmitoylation enriches ER‐localized proteins on the MAM. We demonstrate that palmitoylation of cysteine residue(s) adjacent to the membrane‐spanning domain promotes MAM enrichment of the transmembrane thioredoxin family protein TMX. In addition to TMX, our results also show that calnexin shuttles between the rough ER and the MAM depending on its palmitoylation status. Mutation of the TMX and calnexin palmitoylation sites and chemical interference with palmitoylation disrupt their MAM enrichment. Since ER‐localized heme oxygenase‐1, but not cytosolic GRP75 require palmitoylation to reside on the MAM, our findings identify palmitoylation as key for MAM enrichment of ER membrane proteins.

RAB32 modulates apoptosis onset and mitochondria-associated membrane (MAM) properties

The mitochondria-associated membrane (MAM) has emerged as an endoplasmic reticulum (ER) signaling hub that accommodates ER chaperones, including the lectin calnexin. At the MAM, these chaperones control ER homeostasis but also play a role in the onset of ER stress-mediated apoptosis, likely through the modulation of ER calcium signaling. These opposing roles of MAM-localized chaperones suggest the existence of mechanisms that regulate the composition and the properties of ER membrane domains. Our results now show that the GTPase Rab32 localizes to the ER and mitochondria, and we identify this protein as a regulator of MAM properties. Consistent with such a role, Rab32 modulates ER calcium handling and disrupts the specific enrichment of calnexin on the MAM, while not affecting the ER distribution of protein-disulfide isomerase and mitofusin-2. Furthermore, Rab32 determines the targeting of PKA to mitochondrial and ER membranes and through its overexpression or inactivation increases the phosphorylation of Bad and of Drp1. Through a combination of its functions as a PKA-anchoring protein and a regulator of MAM properties, the activity and expression level of Rab32 determine the speed of apoptosis onset.

Ero1α requires oxidizing and normoxic conditions to localize to the mitochondria-associated membrane (MAM)

Protein secretion from the endoplasmic reticulum (ER) requires the enzymatic activity of chaperones and oxidoreductases that fold polypeptides and form disulfide bonds within newly synthesized proteins. The best-characterized ER redox relay depends on the transfer of oxidizing equivalents from molecular oxygen through ER oxidoreductin 1 (Ero1) and protein disulfide isomerase to nascent polypeptides. The formation of disulfide bonds is, however, not the sole function of ER oxidoreductases, which are also important regulators of ER calcium homeostasis. Given the role of human Ero1α in the regulation of the calcium release by inositol 1,4,5-trisphosphate receptors during the onset of apoptosis, we hypothesized that Ero1α may have a redox-sensitive localization to specific domains of the ER. Our results show that within the ER, Ero1α is almost exclusively found on the mitochondria-associated membrane (MAM). The localization of Ero1α on the MAM is dependent on oxidizing conditions within the ER. Chemical reduction of the ER environment, but not ER stress in general leads to release of Ero1α from the MAM. In addition, the correct localization of Ero1α to the MAM also requires normoxic conditions, but not ongoing oxidative phosphorylation.

Palmitoylation is the switch that assigns calnexin to quality control or ER Ca2+ signaling

Summary The palmitoylation of calnexin serves to enrich calnexin on the mitochondria-associated membrane (MAM). Given a lack of information on the significance of this finding, we have investigated how this endoplasmic reticulum (ER)-internal sorting signal affects the functions of calnexin. Our results demonstrate that palmitoylated calnexin interacts with sarcoendoplasmic reticulum (SR) Ca2+ transport ATPase (SERCA) 2b and that this interaction determines ER Ca2+ content and the regulation of ER–mitochondria Ca2+ crosstalk. In contrast, non-palmitoylated calnexin interacts with the oxidoreductase ERp57 and performs its well-known function in quality control. Interestingly, our results also show that calnexin palmitoylation is an ER-stress-dependent mechanism. Following a short-term ER stress, calnexin quickly becomes less palmitoylated, which shifts its function from the regulation of Ca2+ signaling towards chaperoning and quality control of known substrates. These changes also correlate with a preferential distribution of calnexin to the MAM under resting conditions, or the rough ER and ER quality control compartment (ERQC) following ER stress. Our results have therefore identified the switch that assigns calnexin either to Ca2+ signaling or to protein chaperoning.

Oculomotor apraxia and dilated cardiomyopathy with ataxia syndrome: A case report.

ABSTRACT Dilated cardiomyopathy with ataxia syndrome (DCMA) is a rare mitochondrial condition associated with early onset cardiomyopathy and non-progressive ataxia. The cardiac manifestations may be progressive and often severe, resulting in significant morbidity and mortality. While optic nerve atrophy has been described in patients with DCMA, to our knowledge, there have been no reports of additional ocular phenotypes. We present two related Dariusleut Hutterite patients with documented DCMA syndrome and disorders of ocular motility: poor smooth pursuit and difficulty initiating saccadic eye movements and maintaining target fixation. We thus report the first cases of oculomotor apraxia in DCMA syndrome. By identifying these associated findings early in life, we hope to improve both the clinical diagnostic accuracy and timeliness of intervention in cases of DCMA.

Bilateral uveitis and Usher syndrome: a case report

IntroductionUsher syndrome is a genetically heterogeneous condition and represents the most common cause of inherited combined vision and hearing loss. Deficits manifest as sensorineural hearing loss that typically develops at a young age and retinitis pigmentosa that can lead to peripheral vision loss and night blindness. As a result, this syndrome can have a significant impact on a patient’s quality of life.Previous studies have described an association between Usher syndrome and Fuchs’ heterochromic iridocyclitis, a form of non-granulomatous uveitis that generally presents in a unilateral manner. We present a rare finding of bilateral uveitis and, to the best of our knowledge, the first report of granulomatous uveitis as a feature in a patient with Usher syndrome.Case presentationA 45-year-old Caucasian woman with a known history of retinitis pigmentosa presented to our clinic with suspected Usher syndrome, given her report of long-standing hearing loss. Aside from a mild loss in visual acuity, our patient was otherwise asymptomatic. Visual field testing, audiology and electroretinography findings supported the diagnosis of Usher syndrome. With slit lamp examination she was found to have bilateral keratic precipitates, with large, greasy-white, mutton-fat keratic precipitates on the endothelial surface of her left eye. A thorough work-up that included blood tests and imaging was negative for an alternative cause of her uveitis.ConclusionWe present a rare finding of bilateral uveitis and what we believe to be the first reported instance of mutton-fat keratic precipitates and granulomatous uveitis as a feature in a patient with Usher syndrome. By identifying atypical presentations of the disease, we hope to contribute to the range of ophthalmic conditions that may be seen in association with Usher syndrome.

An analysis of strabismus reoperations in Northern Alberta, Canada from 1995 to 2015

OBJECTIVE To report the reoperation rate in a large group of pediatric and adult strabismus patients over a 21-year period in Northern Alberta, Canada. METHODS A retrospective review of 6177 strabismus surgeries from July 1995 to June 2015 on 5125 pediatric and adult patients was conducted to determine the reoperation rate at a single major referral centre. A set of guidelines was implemented in November 2014 recommending delaying reintervention for at least 12 weeks from the initial surgery, with specific exceptions. RESULTS The historical strabismus reoperation rate over a 21-year period was 15.7%. Of those surgeries requiring reoperation, 77.7% required only 1 reoperation, 17.1% required 2 reoperations, 3.1% required 3 reoperations, and 2.1% required 4 or more reoperations. The mean time between surgeries for patients undergoing reoperation was 2.3 years and the median time was 1.0 years. CONCLUSIONS Our study provides insight into the strabismus reoperation rate and the number of subsequent surgeries performed for patients over a 21-year period. Although preoperative ocular alignment, comorbidities, and the status of the operative eye modify the probability of reoperation, our results from a large cohort of patients provide an impression of the rate of reoperation and may be of benefit in the preoperative counseling of patients. Furthermore, we highlight the scarcity of guidelines for the appropriate timing of reoperation. The implementation of specific guidelines may encourage future investigation into trends in reoperation over time and promote ways to avoid unnecessary surgeries, lower health care costs to stakeholders, and ultimately improve patient care.

A diagnosis of Stevens-Johnson Syndrome (SJS) in a patient presenting with superficial keratitis

Purpose To describe a case of Stevens-Johnson syndrome (SJS) diagnosed in a patient presenting with primarily ocular findings where SJS had not been initially suspected. Observations A 23-year-old female presented with a 2 day history of bilateral eye pain, conjunctival injection, decreased visual acuity, and photophobia in the context of a 4 day history of fever, headache, and sore throat. She was found to have bilateral superficial keratitis and treated for suspected early infectious keratitis secondary to extended contact lens wear. She returned the next day with worsening visual symptoms, a new macular rash over her upper torso, and new ulcerating lesions over her buccal and perioral tissue. The patient was diagnosed with SJS. She was successfully treated using systemic cyclosporine with antibiotics and steroid eye drops. Conclusions and importance Ophthalmologists may be the first physicians to diagnose SJS, a life-threatening condition that can initially present with non-specific viral prodromal symptoms and ocular signs alone. This case emphasizes the importance of considering a patients entire clinical history, especially when the presentation is atypical and the diagnosis is not obviously apparent.