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Mitochondria, central organelles pivotal for eukaryotic cell function, extend their influence beyond ATP production, encompassing roles in apoptosis, calcium signaling, and biosynthesis. Recent studies spotlight two emerging determinants of mitochondrial functionality: intramitochondrial viscosity and sulfur dioxide (SO 2) levels. While optimal mitochondrial viscosity governs molecular diffusion and vital processes like oxidative phosphorylation, aberrations are linked with neurodegenerative conditions, diabetes, and cancer. Similarly, SO 2 , a gaseous signaling molecule, modulates energy pathways and oxidative stress responses; however, imbalances lead to cytotoxic sulfite and bisulfite accumulation, triggering disorders such as cancer and cardiovascular anomalies. Our research focused on development of a dual-channel fluorescent probe, applying electron-withdrawing acceptors within a coumarin dye matrix, facilitating monitoring of mitochondrial viscosity and SO 2 in live cells. This probe distinguishes fluorescence peaks at 650 nm and 558 nm, allowing ratiometric quantification of SO 2 without interference from other sulfur species. Moreover, it enables near-infrared viscosity determination, particularly within mitochondria. The investigation employed theoretical calculations utilizing Density Functional Theory (DFT) methods to ascertain molecular geometries and calculate rotational energies. Notably, the indolium segment of the probe exhibited the lowest rotational energy, quantified at 7.38 kcals/mol. The probe featured heightened mitochondrial viscosity dynamics when contained within HeLa cells subjected to agents like nystatin, monensin, and bacterial lipopolysaccharide (LPS). Overall, our innovative methodology elucidates intricate mitochondrial factors, presenting transformative insights into cellular energetics, redox homeostasis, and therapeutic avenues for mitochondrial-related disorders. [Display omitted] • Development of a new fluorescent probe for real-time monitoring of viscosity and sulfur dioxide (SO 2). • The probe detects sulfur dioxide (SO 2) with ratiometric fluorescence and measures viscosity via near-infrared "turn-on" fluorescence. • The probe can detect viscosity in live cells and Drosophila melanogaster larvae. • Theoretical calculations using Density Functional Theory (DFT) methods validate the efficacy and specificity of the probe design. [ABSTRACT FROM AUTHOR]

Two NAD(P)H-biosensing probes consisting of 1,3,3-trimethyl-3 H -indolium and 3-quinolinium acceptors, linked by thiophene, A , and 3,4-ethylenedioxythiophene, B , bridges are detailed. We synthesized probes C and D , replacing the thiophene connection in probe A with phenyl and 2,1,3-benzothiadiazole units, respectively. Probe E was prepared by substituting probe A 's 3-quinolinium unit with a 1-methylquinoxalin-1-ium unit. Probe solutions are non-fluorescent but in the presence of NADH, exhibit near-infrared fluorescence at 742.1 nm and 727.2 nm for probes A and B , respectively, and generate absorbance signals at 690.6 nm and 685.9 nm. In contrast, probes C and D displayed pronounced interference from NADH fluorescence at 450 nm, whereas probe E exhibited minimal fluorescence alterations in response to NAD(P)H. Pre-treatment of A549 cells with glucose in the presence of probe A led to a significant increase in fluorescence intensity. Additionally, subjecting probe A to lactate and pyruvate molecules resulted in opposite changes in NAD(P)H levels, with lactate causing a substantial increase in fluorescence intensity, conversely, pyruvate resulted in a sharp decrease. Treatment of A549 cells with varying concentrations of the drugs cisplatin, gemcitabine, and camptothecin (5, 10, and 20 µM) led to a concentration-dependent increase in intracellular fluorescence intensity, signifying a rise in NAD(P)H levels. Finally, fruit fly larvae were treated with different concentrations of NADH and cisplatin illustrating applicability to live organisms. The results demonstrated a direct correlation between fluorescence intensity and the concentration of NADH and cisplatin, respectively, further confirming the efficacy of probe A in sensing changes in NAD(P)H levels within a whole organism. [Display omitted] • NAD(P)H-biosensing probes for near-infrared detection. • Reduced background fluorescence, autofluorescence, and cellular damage. • Detect changes in NAD(P)H levels in A549 cells. • Responds to glucose, lactate, pyruvate, and various cancer drugs. • Probe applied to freshly hatched fruit fly larvae. [ABSTRACT FROM AUTHOR]

With the widespread use of avermectins (AVMs) for managing parasitic and agricultural pests, the resistance of worms and insects to AVMs has emerged as a serious threat to human health and agriculture worldwide. The reduced penetration of AVMs is one of the main reasons for the development of the resistance to the chemicals. However, the detailed molecular mechanisms remain elusive. Here, we use the larvae of Drosophila melanogaster as the model organism to explore the molecular mechanisms underlying the development of penetration resistance to AVMs. We clearly show that the chitin layer is thickened and the efflux transporter P-glycoprotein (P-gp) is overexpressed in the AVM-resistant larvae epidermis. We reveal that the activation of the transcription factor Relish by the over-activated epidermal growth factor receptor (EGFR)/AKT/ERK pathway induces the overexpression of the chitin synthases DmeCHS1/2 and P-gp in the resistant larvae. Interestingly, we discover for the first time, to the best of our knowledge, that AVM directly interacts with EGFR and leads to the activation of the EGFR/AKT/ERK pathway, which activates the transcription factor Relish and induces the overexpression of DmeCHS1/2 and P-gp. These findings provide new insights into the molecular mechanisms underlying the development of penetration resistance to drugs.

Navigating across light gradients is essential for survival for many animals. However, we still have a poor understanding of the algorithms that underlie such behaviors. Here, we developed a novel closed-loop phototaxis assay for Drosophila larvae in which light intensity is always spatially uniform but updates depending on the location of the animal in the arena. Even though larvae can only rely on temporal cues during runs, we find that they are capable of finding preferred areas of low light intensity. Further detailed analysis of their behavior reveals that larvae turn more frequently and that heading angle changes increase when they experience brightness increments over extended periods of time. We suggest that temporal integration of brightness change during runs is an important - and so far largely unexplored - element of phototaxis., Competing Interests: Competing interests The authors declare no competing or financial interests., (© 2021. Published by The Company of Biologists Ltd.)

We report a near-infrared fluorescent probe A for the ratiometric detection of cysteine based on FRET from a coumarin donor to a near-infrared rhodamine acceptor. Upon addition of cysteine, the coumarin fluorescence increased dramatically up to 18-fold and the fluorescence of the rhodamine acceptor decreased moderately by 45 % under excitation of the coumarin unit. Probe A has been used to detect cysteine concentration changes in live cells ratiometrically and to visualize fluctuations in cysteine concentrations induced by oxidation stress through treatment with hydrogen peroxide or lipopolysaccharide (LPS). Finally, probe A was successfully applied for the in vivo imaging of Drosophila melanogaster larvae to measure cysteine concentration changes., (© 2019 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.)