Protein homeostasis (proteostasis) disturbances and inflammation are evident in normal aging and some age-related neurodegenerative diseases. While the proteostasis network maintains the integrity of intracellular and extracellular functional proteins, inflammation is a biological response to harmful stimuli. Cellular stress conditions can cause protein damage, thus exacerbating protein misfolding and leading to an eventual overload of the degradation system. The regulation of proteostasis network is particularly important in postmitotic neurons due to their limited regenerative capacity. Therefore, maintaining balanced protein synthesis, handling unfolding, refolding, and degrading misfolded proteins are essential to preserve all cellular functions in the central nervous sysytem. Failing proteostasis may trigger inflammatory responses in glial cells, and the consequent release of inflammatory mediators may lead to disturbances in proteostasis. Here, we review the mechanisms of proteostasis and inflammatory response, emphasizing their role in the pathological hallmarks of neurodegenerative diseases such as Alzheimer’s disease, Parkinson’s disease, and amyotrophic lateral sclerosis. Furthermore, we discuss the interplay between proteostatic stress and excessive immune response that activates inflammation and leads to dysfunctional proteostasis.

Synonymous or silent mutations are usually thought to evolve neutrally. However, accumulating recent evidence has demonstrated that silent mutations may destabilize RNA structures or disrupt cis regulatory motifs superimposed on coding sequences. Such observations suggest the existence of stretches of codon sites that are evolutionary conserved at both DNA-RNA and protein levels. Such stretches may point to functionally important regions within protein coding sequences not necessarily reflecting functional constraints on the amino-acid sequence. The HIV-1 genome is highly compact, and often harbors overlapping functional elements at the protein, RNA, and DNA levels. This superimposition of functions leads to complex selective forces acting on all levels of the genome and proteome. Considering the constraints on HIV-1 to maintain such a highly compact genome, we hypothesized that stretches of synonymous conservation would be common within its genome.

Traditional Western blotting is one of the most used analytical techniques in biological research. However, it can be time-consuming and suffer from a lack of reproducibility. Consequently, devices with different degrees of automation have been developed. These include semi-automated techniques and fully automated devices that replicate all stages downstream of the sample preparation, including sample size separation, immunoblotting, imaging, and analysis. We directly compared traditional Western blotting with two different automated systems, iBind™ Flex, which is a semi-automated system designed to perform the immunoblotting, and JESS Simple Western™, a fully automated and capillary-based system performing all steps downstream of sample preparation and loading, including imaging and image analysis. We found that a fully automated system can save time and importantly offer valuable sensitivity. This is particularly beneficial for limited sample amounts. The downside of automation is the cost of devices and reagents. Nevertheless, automation can be a good option to increase output and facilitate sensitive protein analyses.

Adeno-associated virus (AAV) vectors are currently used in four approved gene therapies for Leber congenital amaurosis (Luxturna), spinal muscular atrophy (Zolgensma), aromatic L-amino acid decarboxylase deficiency (Upstaza) and Haemophilia A (Roctavian), with several more therapies being investigated in clinical trials. AAV gene therapy has long been considered extremely safe both in the context of immunotoxicity and genotoxicity, but recent tragic deaths in the clinical trials for X-linked myotubular myopathy and Duchenne’s muscular dystrophy, together with increasing reports of potential hepatic oncogenicity in animal models have prompted re-evaluation of how much trust we can place on the safety of AAV gene therapy, especially at high doses. In this review we cover genome and capsid engineering strategies that can be used to improve safety of the next generation AAV vectors both in the context of immunogenicity and genotoxicity and discuss the gaps that need filling in our current knowledge about AAV vectors.

Peripheral artery disease (PAD) is a vascular disorder caused by occlusive atherosclerosis, which commonly impairs blood flow to the lower extremities. The prevalence of PAD is increasing globally with >200 million people affected. PAD remains a growing global health problem as the population continues to age and diabetes incidence grows. Many patients with PAD, most notably those with critical limb ischemia, fail attempts at surgical and percutaneous intervention to improve blood flow and are at risk of amputation. Gene therapy provides an opportunity to change the clinical course of PAD in these patients via strategies that increase vascular supply through angiogenesis and arteriogenesis improving muscle perfusion and function in ischemic legs. This article discusses gene therapy approaches in the context of PAD, both intermittent claudication and critical limb ischemia, and the promise of adeno-associated virus–based strategies delivering not just VEGFs (vascular endothelial growth factors) but a range of other mediators as potential new therapeutics. We also highlight challenges and failures in the clinical translation of gene therapy for PAD and how at least some of these obstacles may be overcome using adeno-associated virus.

Significance: The extent and breadth of transcription have recently been uncovered and this has revealed an extensive array of noncoding RNAs (ncRNAs). The biological role and significance of these ncRNAs have been realized and to date it appears that ncRNAs may have many important regulatory functions. ncRNAs are multifaceted and they induce a complexity of different types of transcriptional and posttranscriptional regulation, including gene activation. Recent Advances: Association of ncRNAs with gene activation is an important finding. Not only enhancer RNA (eRNA) but other types of ncRNAs, including small RNA (sRNA), long-noncoding RNA (lncRNA), microRNA (miRNA), and PIWI-associated RNA (piRNA), have also been implicated in gene activation. Interestingly, they often coincide with histone modifications that favor an open chromatin. In addition, these ncRNAs can recruit key factors important for transcription, including RNA polymerase II. They may directly bind the genomic DNA or act as scaffolds; alternatively, they may loop the chromatin to enhance transcription. Critical Issues: Although the role of small activating (sa)RNAs has been considerably studied, the roles of miRNAs and piRNAs in gene activation still need to be substantiated and issues of specificity require further studies. Future Directions: The ncRNA field is coming out of its infancy and we are gaining a global picture of the importance of ncRNAs. However, detailed mechanisms of action of the different ncRNAs are still to be determined. This may reveal novel ways of transcriptional regulation, which will facilitate our ability to utilize these regulatory pathways for research and therapeutic purposes. Antioxid. Redox Signal. 00, 000–000.