James F. Collawn

Key Points: Macrophage accumulation in cystic kidney disease is not directly regulated by the cilia-dependent cyst activation pathway. Macrophage accumulation and cytokine expression are not driving cyst initiation but rather parallel cyst expansion and contribute to cyst progression. Background: Kidney cyst formation occurs due to loss of cilia-localized polycystin proteins ( e.g ., Pkd1 or Pkd2 ) or ciliary structure ( e.g ., Ift88 or Kif3a ). However, cyst progression is more rapid in polycystin mutant mice compared with cilia mutant mice, and loss of cilia in the polycystin mutant background ( e.g ., Pkd2 and Kif3a mutation) greatly attenuates cyst development. This led to the proposal that the polycystins function to repress a cyst-promoting pathway that is dependent on an intact cilium, this is referred to as the cilia-dependent cyst activation pathway. Renal macrophages are also involved in regulating cyst progression, but it is unknown whether this occurs through the cilia-dependent cyst activation or separate pathway. Methods: To examine whether macrophage accumulation was regulated through a cilia-dependent pathway, we compared macrophage accumulation and cytokine expression levels in Pkd2 mutant kidneys with or without intact cilia ( Ift88 mutants). To avoid the impact of cyst-induced damage on macrophage accumulation, we conducted comparisons after standardizing the samples for cystic indices between the Pkd2 , Ift88 , and Pkd2;Ift88 double mutants. Results: Disruption of Ift88 in Pkd2 mutants reduced cyst burden and attenuated macrophage accumulation and cytokine expression levels. However, when the mutants were standardized based on cystic indices, no significant differences in macrophage number or cytokine expression were evident between the Pkd2 , Ift88 , and Pkd2;Ift88 double mutants at either early or advanced stage of cyst progression, and pathway analysis revealed that the macrophage populations were similar between groups based on single-cell RNAseq data. Conclusions: These data indicated that macrophage accumulation and cytokine expression did not drive cyst initiation but rather paralleled cyst expansion regardless of the genotype or rate of disease progression through a cilia-independent pathway.

Ubiquitin, a small protein, is well known for tagging target proteins through a cascade of enzymatic reactions that lead to protein degradation. The ubiquitin tag, apart from its signaling role, is paramount in destabilizing the modified protein. Here, we explore the complex role of ubiquitin-mediated protein destabilization in the intricate proteolysis process by the 26S proteasome. In addition, the significance of the so-called ubiquitin-independent pathway and the role of the 20S proteasome are considered. Next, we discuss the ubiquitin–proteasome system’s interplay with pathogenic microorganisms and how the microorganisms manipulate this system to establish infection by a range of elaborate pathways to evade or counteract host responses. Finally, we focus on the mechanisms that rely either on (i) hijacking the host and on delivering pathogenic E3 ligases and deubiquitinases that promote the degradation of host proteins, or (ii) counteracting host responses through the stabilization of pathogenic effector proteins.

Efficient brain function requires as much as 20% of the total oxygen intake to support normal neuronal cell function. This level of oxygen usage, however, leads to the generation of free radicals, and thus can lead to oxidative stress and potentially to age-related cognitive decay and even neurodegenerative diseases. The regulation of this system requires a complex monitoring network to maintain proper oxygen homeostasis. Furthermore, the high content of mitochondria in the brain has elevated glucose demands, and thus requires a normal redox balance. Maintaining this is mediated by adaptive stress response pathways that permit cells to survive oxidative stress and to minimize cellular damage. These stress pathways rely on the proper function of the endoplasmic reticulum (ER) and the activation of the unfolded protein response (UPR), a cellular pathway responsible for normal ER function and cell survival. Interestingly, the UPR has two opposing signaling pathways, one that promotes cell survival and one that induces apoptosis. In this narrative review, we discuss the opposing roles of the UPR signaling pathways and how a better understanding of these stress pathways could potentially allow for the development of effective strategies to prevent age-related cognitive decay as well as treat neurodegenerative diseases.

MicroRNAs (miRNAs) play a critical role in the regulation of mRNA stability and translation. In spite of our present knowledge on the mechanisms of mRNA regulation by miRNAs, the utilization and translation of these ncRNAs into clinical applications have been problematic. Using hsa-miR-429 as an example, we discuss the limitations encountered in the development of efficient miRNA-related therapies and diagnostic approaches. The miR-200 family members, which include hsa-miR-429, have been shown to be dysregulated in different types of cancer. Although these miR-200 family members have been shown to function in suppressing epithelial-to-mesenchymal transition, tumor metastasis, and chemoresistance, the experimental results have often been contradictory. These complications involve not only the complex networks involving these noncoding RNAs, but also the problem of identifying false positives. To overcome these limitations, a more comprehensive research strategy is needed to increase our understanding of the mechanisms underlying their biological role in mRNA regulation. Here, we provide a literature analysis of the verified hsa-miR-429 targets in various human research models. A meta-analysis of this work is presented to provide better insights into the role of hsa-miR-429 in cancer diagnosis and any potential therapeutic approach.

Despite our understanding of the unfolded protein response (UPR) pathways, the crosstalk between the UPR and the complex signaling networks that different cancers utilize for cell survival remains to be, in most cases, a difficult research barrier. A major problem is the constant variability of different cancer types and the different stages of cancer as well as the complexity of the tumor microenvironments (TME). This complexity often leads to apparently contradictory results. Furthermore, the majority of the studies that have been conducted have utilized two-dimensional in vitro cultures of cancer cells that were exposed to continuous hypoxia, and this approach may not mimic the dynamic and cyclic conditions that are found in solid tumors. Here, we discuss the role of intermittent hypoxia, one of inducers of the UPR in the cellular component of TME, and the way in which intermittent hypoxia induces high levels of reactive oxygen species, the activation of the UPR, and the way in which cancer cells modulate the UPR to aid in their survival. Although the past decade has resulted in defining the complex, novel non-coding RNA-based regulatory networks that modulate the means by which hypoxia influences the UPR, we are now just to beginning to understand some of the connections between hypoxia, the UPR, and the TME.

The cellular adaptive response to hypoxia relies on the expression of hypoxia‐inducible factors (HIFs), HIF‐1 and HIF‐2. HIFs regulate global gene expression changes during hypoxia that are necessary for restoring oxygen homeostasis and promoting cell survival. In the early stages of hypoxia, HIF‐1 is elevated, whereas at the later stages, HIF‐2 becomes the predominant form. What governs the transition between the two HIFs (the HIF switch) and the role of miRNAs in this regulation are not completely clear. Genome‐wide expression studies on the miRNA content of RNA‐induced silencing complexes (RISC) in HUVECs exposed to hypoxia compared to the global miRNA‐Seq analysis revealed very specific differences between these two populations. We analyzed the miRNA and mRNA composition of RISC at 2 h (mainly HIF‐1 driven), 8 h (HIF‐1 and HIF‐2 elevated), and 16 h (mainly HIF‐2 driven) in a gene ontology context. This allowed for determining the direct impact of the miRNAs in modulating the cellular signaling pathways involved in the hypoxic adaptive response. Our results indicate that the miRNA‐mRNA RISC components control the adaptive responses, and this does not always rely on the miRNA transcriptional elevations during hypoxia. Furthermore, we demonstrate that the hypoxic levels of the vast majority of HIF‐1‐dependent miRNAs (including miR‐210‐3p) are also HIF‐2 dependent and that HIF‐2 governs the expression of 11 specific miRNAs. In summary, the switch from HIF‐1 to HIF‐2 during hypoxia provides an important level of miRNA‐driven control in the adaptive pathways in endothelial cells.

The adaptive response to hypoxia involves the transcriptional induction of three transcription factors called hypoxia inducible factor alpha 1, 2 and 3 (HIF-1α, HIF-2α, and HIF-3α) which dimerize with constitutively expressed beta chains that together form the HIF-1, -2 and -3 transcription factors. During normoxic conditions, the alpha chain is expressed at low levels since its stability is regulated by prolyl-hydroxylation that promotes subsequent ubiquitination and degradation. During hypoxic conditions, however, the prolyl hydroxylases are less active, and the alpha chain accumulates through elevated protein stability and the elevated induction of expression. Two of the three HIFs isoforms present in mammals, HIF-1 and HIF-2, are well characterized and have overlapping functions that promote cell survival, whereas HIF-3’s role remains less clear. The HIF-3 response is complicated because the HIF3A gene can utilize different promotors and alternate splicing sites that result in a number of different HIF-3α isoforms. Here, using human umbilical vein endothelial cells (HUVECs), we demonstrate that one of the isoforms of HIF-3α, isoform 2 (HIF-3α2) accumulates at a late stage of hypoxia and induces the expression of DNA damage inducible transcript 3 (DDIT4), a gene known to promote apoptosis. We also demonstrate that caspase 3/7 activity is elevated, supporting that the role of the HIF-3α2 isoform is to promote apoptosis. Furthermore, we provide evidence that HIF-3α2 is also expressed in seven other primary endothelial cell types, suggesting that this may be a common feature of HIF-3α2 in endothelial cells.

Tremendous progress in RNAi delivery methods and design has allowed for the effective development of siRNA-based therapeutics that are currently under clinical investigation for various cancer treatments. This approach has the potential to revolutionize cancer therapy by providing the ability to specifically downregulate or upregulate the mRNA of any protein of interest. This exquisite specificity, unfortunately, also has a downside. Genetic variations in the human population are common because of the presence of single nucleotide polymorphisms (SNPs). SNPs lead to synonymous and non-synonymous changes and they occur once in every 300 base pairs in both coding and non-coding regions in the human genome. Much less common are the somatic mosaicism variations associated with genetically distinct populations of cells within an individual that is derived from postzygotic mutations. These heterogeneities in the population can affect the RNAi’s efficacy or more problematically, which can lead to unpredictable and sometimes adverse side effects. From a more positive viewpoint, both SNPs and somatic mosaicisms have also been implicated in human diseases, including cancer, and these specific changes could offer the ability to effectively and, more importantly, selectively target the cancer cells. In this review, we discuss how SNPs in the human population can influence the development and success of novel anticancer RNAi therapies and the importance of why SNPs should be carefully considered.

Cold viruses have generally been considered fairly innocuous until the appearance of the severe acute respiratory coronavirus 2 (SARS-CoV-2) in 2019, which caused the coronavirus disease 2019 (COVID-19) global pandemic. Two previous viruses foreshadowed that a coronavirus could potentially have devastating consequences in 2002 [severe acute respiratory coronavirus (SARS-CoV)] and in 2012 [Middle East respiratory syndrome coronavirus (MERS-CoV)]. The question that arises is why these viruses are so different from the relatively harmless cold viruses. On the basis of an analysis of the current literature and using bioinformatic approaches, we examined the potential human miRNA interactions with the SARS-CoV-2’s genome and compared the miRNA target sites in seven coronavirus genomes that include SARS-CoV-2, MERS-CoV, SARS-CoV, and four nonpathogenic coronaviruses. Here, we discuss the possibility that pathogenic human coronaviruses, including SARS-CoV-2, could modulate host miRNA levels by acting as miRNA sponges to facilitate viral replication and/or to avoid immune responses.