Title : Cadmium Selenide Zinc Sulfide Quantum Dots on Saccharomyces cerevisiae: Their traffic routes and impact on endocytosis and exocytosis as well as on actin dynamics
Abstract:
Quantum dots (QDs) are nano-sized semiconductor crystals that are highly utilized for research and medical purposes. Although recent studies have hinted at the toxicity of QDs, their impact on fungal cells remains unclear. Instead of relying on each biochemical assay to assess the harmful effects mediated by QDs, we have used RNAseq to create altered transcriptome profiles in response to QD treatment. We found that cadmium-based QDs leads to an alteration in the quantity of mRNA transcripts implicated in endocytosis, membrane fusion, sorting at the endosome, and endosome maturation. We then examined the trafficking route of red Cadmium Selenide Zinc Sulfide Quantum Dots (CdSe/ZnS QDs) as well as the phenotypic changes it induces in budding yeast cells. We tracked CdSe/ZnS QDs’ subcellular location by using yeast strains expressing different reference markers, including: plasma membrane reference marker GFP-2PH; early endocytosis vesicle reference marker Abp1- GFP; late Golgi/trans Golgi network reference marker FAPPI(PH)-GFP; and late endosome reference marker Vps10-GFP. We found that immediately after treating, CdSe/ZnS QDs interacted with yeast cells. However, CdSe/ZnS QD only co-localized with the plasma membrane reference marker after 3 hours of treatment, suggesting that CdSe/ZnS requires at least 3hrs to arrive at the plasma membrane. Around 6 hours after treatment, CdSe/ZnS was found at the plasma membrane, the early endocytosis vesicle, and the late Golgi/ trans Golgi network. QDs was never found co-localizing with the late endosome, suggesting that this is not a destination for CdSe/ZnS QDs intracellular trafficking. The rest of QDs’ intracellular trafficking route is in need of further investigation. We also treated yeast with different concentrations of CdSe/ZnS QDs (4µg/mL, 12µg/mL, 50µg/mL) and studied the integrity of actin cable upon QDs exposure. After 6 hours, a higher percentage of cells showed actin cable fragmentation in 12µg/mL and 50µg/mL of QDs treatment. Furthermore, we performed a recovery assay by removing QDs from the culture media. After 3 hours of incubation in QDs- free media, partial actin cable recovery was observed, hinting that the effect of QDs toxicity is reversible. Additionally, our viability assay data reveals that 6hrs of QDs exposure led to a significant decrease in the yeast sample’s optical density for all treatment concentrations. Interestingly, there was no significant difference in optical density between the treated samples and the control samples after 24 hours of treatment, indicating that QDs exposure only causes an inhibitory effect on yeast growth. We are currently assessing if QDs affect the membrane cargo Snc1 trafficking. Altogether, our research will provide new insights into the understanding of QD-mediated impacts on endocytosis and exocytosis.
Audience take-away:
- Quantum dot chemical and physical properties, Quantum-dot impacts on the alteration of transcriptome, Transcriptome data-based hypothesis would be tested for more investigation on physiological impacts mediated by QDs.
- Over the last five years, we have characterized the toxicity of several Engineered Nanomaterials (ENMs) that are not strictly regulated by the FDA or EPA in the United States. ENMs, being nano- sized particles (1-100 nm in diameter), have a significant number of applications and are currently used in healthcare as fluorescent probes and drug delivery vehicles.
- They are also used in consumer products including textiles, cosmetics, and food. According to the EPA’s Information Gathering Rule, companies must report on the chemical identity, manufacturing methods, and exposure of any nanoscale materials they incorporate into their products. We provided gene expression data in yeast and mammalian cells exposed to nanoparticles, including carbon nanotubes, silver nanoparticles, metal oxide nanoparticles (ZnO and CuO), and quantum dots (CdSe/ZnS and InP/ZnS). With our results, we have identified thousands of genes and many cellular functions that were negatively impacted by ENM exposure.
- Notably, our contributions provided novel insights into the field of ENM-toxicity by implementing state-of-the-art transcriptomic techniques (RNA- seq). Although these agencies have attempted to manage the risks of ENM exposure, there are legal frameworks preventing their regulation because of their poorly understood toxicity. Currently, we have a poor understanding of ENM-toxicity, however, we continue to use them.
- These reasons are precisely why new government regulations and oversight are needed to responsibly use them. A big step in updating the current ENM government regulations is conducting a thorough investigation of their toxicity. Our findings on ENMs have ultimately helped us to understand how they are toxic through identifying changes in gene expression and what functions those genes are implicated in. Eventually, when we better understand how these extremely small materials behave, we can improve upon current regulation policies that will more efficiently protect public health and the environment.
