A new method for senescent cell detection is described, which is based on lipofuscin labeling with a fluorescent reporter through a biorthogonal strain-promoted azide-alkyne cycloaddition. The sensing protocol involves a first step where the interaction of lipofuscin with a Sudan Black B derivative containing an azide moiety (SBB-N3 ) is carried out. In the final step, the azide moiety reacts with a fluorophore containing a cyclooctene ring (BODIPY). The efficacy of this two-step protocol is assessed in senescent melanoma SK-MEL-103 cells, senescent triple-negative breast cancer MDA-MB-231 cells and senescent WI-38 fibroblasts. In all cases, a clear fluorescence pattern was observed in senescent cells, compared to proliferative cells, only when the SBB-N3 -BODIPY probe was formed. Our results provide an alternative tool for the detection of senescent cells, based on an in situ bio-orthogonal reaction for lipofuscin labeling.

The haematopoietic system is maintained by rare haematopoietic stem cells (HSCs), which are quiescent most of the time and only divide occasionally to self-renew and/or to undergo commitment to clonal expansion via the generation of highly proliferative progenitor cells. The latter is responsible for the generation of all mature cells of the system through subsequent lineage commitment and terminal differentiation. Cells with similar properties also exist in leukaemias and are known as leukaemia stem cells (LSCs). Quiescence provides essential protection for both HSC and LSC from cytotoxic stress and DNA damage and, in the case of LSCs, likely causes therapy resistance and disease relapse in leukaemia patients. Specific inhibition of LSC quiescence has been considered a promising strategy for eliminating LSCs and curing leukaemias. Although the understanding of mechanisms responsible for quiescence maintenance in these cells remains limited, particularly for LSCs, recent studies have suggested potential differences in their dependency on certain pathways and their levels of stress and DNA damage caused by increased cycling. Such differences likely stem from oncogenic mutations in LSCs and could be specifically exploited for the elimination of LSCs while sparing normal HSCs in the future.

It has been recently shown that in some proteins, tertiary-structure dynamics occur surprisingly fast, that is on the microsecond or sub-millisecond time scales. In this State of the Art Review, we discuss how such ultrafast domain motions relate to the function of caseinolytic peptidase B (ClpB), a AAA+ disaggregation machine. ClpB is a large hexameric protein that collaborates with cellular chaperone machinery to rescue protein chains from aggregates. We used single-molecule FRET spectroscopy to capture the dynamics of essential structural elements within this machine. It was found that the middle domain of ClpB, known to act as its activator, toggles between two states much faster than the overall activity cycle of the protein, suggesting a novel mode of continuous, tunable switching. Motions of the N-terminal domain were observed to restrict the conformational space of the M domain in the absence of a substrate protein, thereby preventing it from tilting and spuriously activating ClpB. Finally, microsecond dynamics of pore loops responsible for substrate pulling through ClpB's central channel, together with their response to specific perturbations, point to a Brownian-ratchet mechanism for protein translocation. Based on our findings, we propose a two-time-scale model for the activity of ClpB, in which fast conformational dynamics affect slower functional steps, determined by ATP hydrolysis time. Future work on this and other proteins is likely to shed further light on the role of ultrafast dynamics on protein function.

Cilia are microtubule-based organelles that protrude from the cell surface and play crucial roles in cellular signaling pathways and extracellular fluid movement. Defects in the ciliary structures and functions are implicated in a set of hereditary disorders, including polycystic kidney disease, nephronophthisis, and Bardet-Biedl syndrome, which are collectively termed as ciliopathies. The application of mass spectrometry-based proteomic approaches to explore ciliary components provides important clues for understanding their physiological and pathological roles. In this review, we focus primarily on proteomic studies involving the identification of proteins in motile cilia and primary cilia, proteomes in ciliopathies, and interactomes of ciliopathy proteins. Collectively, the integration of these data sets will be beneficial for the comprehensive understanding of ciliary structures and exploring potential biomarkers and therapeutic targets for ciliopathies.

The central nervous system is particularly susceptible to DNA repair deficiency, which renders a variety of neurodevelopmental and neurodegenerative disorders in humans. It is generally believed that DNA damage occurs upon repetitive replication and oxidative stress in highly proliferating neuroprogenitor cells (NPs), or due to high rates of metabolism and active neuronal activity in terminally differentiated neurons. DNA double-stranded breaks (DSBs) and single-stranded breaks (SSBs) constitute the most prevalent forms of DNA damage, which can result in neuronal apoptosis if unrepaired. Despite these notions, there are still gaps in our knowledge regarding the mechanism and specificity of DNA damage and repair in the neural development and the homeostasis of neural tissues. Recent studies have identified recurrent DSBs within neuronal long genes in NPs and 'programmed' SSBs in neuronal activity genes. However, the physiological function of these DNA breakages in the nervous system has not been so far explored. In this review, we summarise the recent advances in the field of DNA damage and DNA repair in neural development and neuropathies.

Transcriptome-wide interrogation of eukaryotic genomes has unveiled the pervasive nature of RNA polymerase II transcription. Virtually, any DNA region with an accessible chromatin structure can be transcribed, resulting in a mass production of noncoding RNAs (ncRNAs) with the potential of interfering with gene expression programs. Budding yeast has proved to be a powerful model organism to understand the mechanisms at play to control pervasive transcription and overcome the risks of hazardous disruption of cellular functions. In this review, we focus on the actors and strategies yeasts employ to govern ncRNA production, and we discuss recent findings highlighting the dangers of losing control over pervasive transcription.

At the time of writing, there were 486 761 597 global cases of COVID-19 with 6 142 735 confirmed deaths (World Health Organization, 4 April 2022). According to the scarcity of information about estimation of cases with mild or no symptoms, it is suggested that they could represent 25-80% of all infections. The majority of these cases remain untested, although they are infective. The molecular diagnosis of COVID-19 is based mainly on quantitative reverse transcription PCR. However, this approach faces several challenges related to the shortage of resources and people who are adequately trained to run the tests. Alternative testing methods, targeting effectively several viral compounds at different stages of the infection, have quickly emerged. However, universal systems that are specific, sensitive, affordable, easy, portable and scalable are still warranted. In this review, a comprehensive compilation of the methods available is provided.