Downregulation of the circadian Period family genes is positively correlated with poor head and neck squamous cell carcinoma prognosis

ABSTRACT

We investigated the relationship between head and neck squamous cell carcinoma (HNSCC) and the mRNA and protein expression levels of the circadian genes of the Period (Per) family, Per1, Per2 and Per3. Tissue sections of HNSCC and normal head and neck tissues from two patient cohorts from two different hospitals were collected to assess the mRNA and protein expressions of the three Per family genes using real-time quantitative PCR (RT-PCR) and immunohistochemistry (IHC). The clinicopathological features and disease prognosis for the latter cohort were analyzed through IHC and statistical methods. Protein positive expression levels of the three Per family genes in HNSCC tissues was found to be approximately two times lower than that in normal tissues (p < .01). Moreover, patients with locally advanced HNSCC showed significantly greater downregulation of Per1, Per2 and Per3 mRNA expression levels as compared to patients with early-stage cancer (p < .05). Immunohistochemical examination of HNSCC patient tissues revealed a positive correlation between the Per family protein expression and the clinical tumor staging (p < .05). In addition, the Per protein-positive expression group showed higher 3-year survival rates [overall survival (OS) and progression-free survival (PFS)] as assessed by Kaplan-Meier plots and statistical analysis (p < .05). Our findings confirm the positive correlation between Per family gene expression and survival outcomes and support their role as prognostic markers for HNSCC.     

Protein interaction affinity determination by quantitative FRET technology

The dissociation constant, K_d, is an important parameter for characterizing protein–protein interaction affinities. SUMOylation is one of the important protein post‐translational modifications and it involves a multi‐step enzymatic cascade reaction, resulting in peptide activation and substrate conjugation. Multiple covalent and non‐covalent protein–protein interactions are involved in this cascade. Techniques involving Förster resonance energy transfer (FRET) have been widely used in biological studies in vitro and in vivo, and they are very powerful tools for elucidating protein interactions in many regulatory cascades. In our previous studies, we reported the attempt to develop a new method for the determination of the K_d by FRET assay using the interaction of SUMO1 and its E2 ligase, Ubc9 as a test system. However, the generality and specifications of this new method have not been fully determined. Here we report a systematic approach for determining the dissociation constant (K_d) in the SUMOylation cascade and for further sensitivity and accuracy testing by the FRET technology. From a FRET donor to acceptor concentration ratio range of 4–40, the K_ds of SUMO1 and Ubc9 consistently agree well with values from surface plasmon resonance and isothermal titration calorimetry. These results demonstrate the high sensitivity and accuracy of the FRET‐based K_d determination approach. This technology, therefore, can be used in general for protein–protein interaction dissociation constant determination. Biotechnol. Bioeng. 2012; 109: 2875–2883. © 2012 Wiley Periodicals, Inc.     

Mind the gap: the minimal detectable separation distance between two objects during active electrolocation

In a food‐rewarded two‐alternative forced‐choice procedure, it was determined how well the weakly electric elephantnose fish Gnathonemus petersii can sense gaps between two objects, some of which were placed in front of complex backgrounds. The results show that at close distances, G. petersii is able to detect gaps between two small metal cubes (2 cm × 2 cm × 2 cm) down to a width of c. 1·5 mm. When larger objects (3 cm × 3 cm × 3 cm) were used, gaps with a width of 2–3 mm could still be detected. Discrimination performance was better (c. 1 mm gap size) when the objects were placed in front of a moving background consisting of plastic stripes or plant leaves, indicating that movement in the environment plays an important role for object identification. In addition, the smallest gap size that could be detected at increasing distances was determined. A linear relationship between object distance and gap size existed. Minimal detectable gap sizes increased from c. 1·5 mm at a distance of 1 cm, to 20 mm at a distance of 7 cm. Measurements and simulations of the electric stimuli occurring during gap detection revealed that the electric images of two close objects influence each other and superimpose. A large gap of 20 mm between two objects induced two clearly separated peaks in the electric image, while a 2 mm gap caused just a slight indentation in the image. Therefore, the fusion of electric images limits spatial resolution during active electrolocation. Relative movements either between the fish and the objects or between object and background might improve spatial resolution by accentuating the fine details of the electric images.