Deletion of prominin-1 in mice results in disrupted photoreceptor outer segment protein homeostasis.

AIM: To illustrate the underlying mechanism how prominin-1 (also known as Prom1) mutation contribute to progressive photoreceptor degeneration. METHODS: A CRISPR-mediated Prom1 knockout (Prom1-KO) mice model in the C57BL/6 was generated and the photoreceptor degeneration phenotypes by means of structural and functional tests were demonstrated. Immunohistochemistry and immunoblot analysis were performed to reveal the localization and quantity of related outer segment (OS) proteins. RESULTS: The Prom1-KO mice developed the photoreceptor degeneration phenotype including the decreased outer nuclear layer (ONL) thickness and compromised electroretinogram amplitude. Immunohistochemistry analysis revealed impaired trafficking of photoreceptor OS proteins. Immunoblot data demonstrated decreased photoreceptor OS proteins. CONCLUSION: Prom1 deprivation causes progressive photoreceptor degeneration. Prom1 is essential for maintaining normal trafficking and normal quantity of photoreceptor OS proteins. The new light is shed on the pathogenic mechanism underlying photoreceptor degeneration caused by Prom1 mutation.

Myofibril diameter is set by a finely tuned mechanism of protein oligomerization in Drosophila.

Myofibrils are huge cytoskeletal assemblies embedded in the cytosol of muscle cells. They consist of arrays of sarcomeres, the smallest contractile unit of muscles. Within a muscle type, myofibril diameter is highly invariant and contributes to its physiological properties, yet little is known about the underlying mechanisms setting myofibril diameter. Here we show that the PDZ and LIM domain protein Zasp, a structural component of Z-discs, mediates Z-disc and thereby myofibril growth through protein oligomerization. Oligomerization is induced by an interaction of its ZM domain with LIM domains. Oligomerization is terminated upon upregulation of shorter Zasp isoforms which lack LIM domains at later developmental stages. The balance between these two isoforms, which we call growing and blocking isoforms sets the stereotyped diameter of myofibrils. If blocking isoforms dominate, myofibrils become smaller. If growing isoforms dominate, myofibrils and Z-discs enlarge, eventually resulting in large pathological aggregates that disrupt muscle function.

Different Evolutionary Trajectories of Two Insect-Specific Paralogous Proteins Involved in Stabilizing Muscle Myofibrils.

Alp/Enigma family members have a unique PDZ domain followed by zero to four LIM domains, and are essential for myofibril assembly across all species analyzed so far. Drosophila melanogaster has three Alp/Enigma family members, Zasp52, Zasp66, and Zasp67. Ortholog search and phylogenetic tree analysis suggest that Zasp genes have a common ancestor, and that Zasp66 and Zasp67 arose by duplication in insects. While Zasp66 has a conserved domain structure across orthologs, Zasp67 domains and lengths are highly variable. In flies, Zasp67 appears to be expressed only in indirect flight muscles, where it colocalizes with Zasp52 at Z-discs. We generated a CRISPR null mutant of Zasp67, which is viable but flightless. We can rescue all phenotypes by re-expressing a Zasp67 transgene at endogenous levels. Zasp67 mutants show extended and broken Z-discs in adult flies, indicating that the protein helps stabilize the highly regular myofibrils of indirect flight muscles. In contrast, a Zasp66 CRISPR null mutant has limited viability, but only mild indirect flight muscle defects illustrating the diverging evolutionary paths these two paralogous genes have taken since they arose by duplication.

Surgical Outcomes of 27-Gauge Pars PLana Vitrectomy for Symptomatic Vitreous Floaters.

PURPOSE: To report the surgical outcomes of 27-gauge pars plana vitrectomy (PPV) for symptomatic vitreous floaters. METHODS: 47 eyes of 47 patients (39 males, 83.0%) with symptomatic vitreous floaters who underwent 27-gauge PPV and followed up for more than 6 months were included. The mean age was 34.7 ± 13.5 years. RESULTS: No operative complication occurred. At first day postoperatively, the intraocular pressure (IOP) was significantly lower than that at other time points (8.6 ± 2.7 mmHg, p < 0.001). 28 (59.6%) eyes had transient hypotony (IOP < 8 mmHg). All were recovered within 1 week postoperatively. The BCVA of 41 eyes (41/47, 87.2%) remained unchanged or improved. Postoperative complications occurred in two eyes: one (2.1%) had endophthalmitis and one (2.1%) had retinal detachment. No clinical significant cataract was observed in the 42 postoperative phakic eyes. 91.5% of the patients were satisfied with the surgery outcome. Besides, 91.3% of the patients felt that the floaters were removed completely or only had an acceptable residual. CONCLUSION: Visual acuity of most patients remained unchanged or improved following 27-gague pars plana vitrectomy for symptomatic vitreous floaters, resulting in high patient satisfaction. However, this treatment should be performed with great caution since severe postoperative complications may still occur. This trial is registered with NCT03049163.

Rapid IFM Dissection for Visualizing Fluorescently Tagged Sarcomeric Proteins.

Sarcomeres, the smallest contractile unit of muscles, are arguably the most impressive actomyosin structure. Yet a complete understanding of sarcomere formation and maintenance is missing. The Drosophila indirect flight muscle (IFM) has proven to be a very valuable model to study sarcomeres. Here, we present a protocol for the rapid dissection of IFM and analysis of sarcomeres using fluorescently tagged proteins.