Surface modifications of carbon nanodots reveal the chemical source of their bright fluorescence.

Fluorescent carbon nanodots (CNDs) have drawn increasing attention in recent years. These cost-effective and eco-friendly nanomaterials with bright fluorescence have been investigated as promising materials for electrooptic and bioimaging applications. However, the chemical source stimulating their strong fluorescence has not been completely identified to date. Depending on the chemical composition, two absorption peaks are observed in the visible range. In this study, we applied selected chemical modifications to CNDs in order to elucidate the correlation between the chemical structure and optical behavior of CNDs. Varying the amount of acetic acid in the synthesis process resulted in different effects on the absorbance and fluorescence photo-spectra. Specifically, at a low concentration (10%), the fluorescence is dramatically red shifted from 340 to 405 nm. Comprehensive characterization of the chemical modification by FTIR and XPS allows identification of the role of acetic acid in the reaction mechanism leading to the modified photoactivity. The functional group responsible for the 405 nm peak was identified as HPPT. We describe a chemical mechanism involving acetic acid that leads to an increased concentration of HPPT groups on the surface of the CNDs. Applying two additional independent chemical and consequently optical modifications namely solution pH and annealing on the nanodots further supports our proposed explanation. Understanding the molecular origin of CND fluorescence may promote the design and control of effective CND fluorescence in optical applications.

Linking protein kinase C to the cell cycle: ectopic expression of PKC eta in NIH3T3 cells alters the expression of cyclins and Cdk inhibitors and induces adipogenesis.

Protein kinase C encodes a family of enzymes implicated in cellular differentiation, growth control and tumor promotion. However, very little is known with respect to the molecular mechanisms that link protein kinase C to cell cycle control. Here we report that ectopic expression of PKC eta in NIH3T3 fibroblasts blocks the normal phosphorylation of the Rb protein in quiescent cultures restimulated to enter the cell cycle; PKC eta activates a cellular program that includes increased expression of cyclins E (but not cyclin D), as well as the induced expression of the cyclin-dependent kinase inhibitors p21WAF1 and p27KIP1. The increased expression of the latter inhibitors and their association with the cyclin E-Cdk2 complex results in decreased cyclin E associated kinase activity. Furthermore, in contrast to the control NIH3T3 cells, the cell that express PKC eta can be induced to undergo adipocyte differentiation in response to adipogenic hormones. Thus, PKC eta induces altered expression of several cell cycle related functions, which may contribute to its ability to promote cellular differentiation.

Molecular modeling based mutagenesis defines ligand binding and specificity determining regions of fibroblast growth factor receptors.

The fibroblast growth factor receptor 2 (FGFR2) and the keratinocyte growth factor receptor (KGFR) have different ligand binding specificities despite differing only in the second half of their immunoglobulin-like (Ig-like) domain III. Three-dimensional model structures were generated for domain III on the basis of variable (V) Ig domains. The region that differs between the two receptors is predicted to include two loops: one connects beta-strands F-G and is analogous to the complementarity determining region 3 (CDR3) of immunoglobulins; the other connects beta-strands D-E. These regions were targeted for mutagenesis. Single mutations in the F-G loop were found to only slightly alter ligand binding, whereas a double mutant, KGFR Y345-->S,Q348-->I, acquired significant affinity for bFGF. Notably, the affinity of this double mutant KGFR for KGF and aFGF was essentially unaltered. A mutant FGFR2, in which the D-E beta-hairpin (T319TDKEI) is replaced with the KGFR D-E beta-hairpin (S319SNA), has 9-fold reduced affinity for bFGF. These results demonstrate that the F-G or CDR3 analogous loop in FGFRs plays a key role in determining ligand binding and specificity. In addition, however, the protein loop connecting beta-strands D and E may also be involved in ligand binding. Several point mutations in FGFR2, shown recently to give rise to multiple inherited skeletal defects, are localized according to our models to the F-G or D-E loops of domain III. Our results strongly suggest that these naturally occurring mutations specifically alter ligand binding by FGFR2.

The protein kinase C-related PKC-L(eta) gene product is localized in the cell nucleus.

The tumor promoters phorbol esters are thought to induce changes in cell growth and gene expression by direct activation of protein kinase C (PKC). However, the molecular mechanisms by which PKC molecules transduce signals into the cell nucleus are unknown. In this study, we provide evidence for a direct target for phorbol esters in the nucleus. We demonstrate that the new PKC-related family member, PKC-L, recently isolated by us, is expressed specifically in the cell nucleus. Localization of PKC-L in the cell nucleus is shown both by immunofluorescence staining and by subcellular fractionation experiments of several human cell lines, including the human epidermoid carcinoma line A431. Treatment of these cells by phorbol esters does not induce the down-regulation of PKC-L, in contrast to their effect on classical PKC family members. This is the only PKC isoenzyme described so far that resides permanently and specifically in the cell nucleus. PKC-L may function as an important link in tumor promoting, e.g., as a nuclear regulator of gene expression that changes the phosphorylation state of transcriptional components such as the AP-1 complex.