Reproducibility efforts as a teaching tool: A pilot study.

The "replication crisis" is a methodological problem in which many scientific research findings have been difficult or impossible to replicate. Because the reproducibility of empirical results is an essential aspect of the scientific method, such failures endanger the credibility of theories based on them and possibly significant portions of scientific knowledge. An instance of the replication crisis, analytic replication, pertains to reproducing published results through computational reanalysis of the authors' original data. However, direct replications are costly, time-consuming, and unrewarded in today's publishing standards. We propose that bioinformatics and computational biology students replicate recent discoveries as part of their curriculum. Considering the above, we performed a pilot study in one of the graduate-level courses we developed and taught at our University. The course is entitled Intro to R Programming and is meant for students in our Master's and PhD programs who have little to no programming skills. As the course emphasized real-world data analysis, we thought it would be an appropriate setting to carry out this study. The primary objective was to expose the students to real biological data analysis problems. These include locating and downloading the needed datasets, understanding any underlying conventions and annotations, understanding the analytical methods, and regenerating multiple graphs from their assigned article. The secondary goal was to determine whether the assigned articles contained sufficient information for a graduate-level student to replicate its figures. Overall, the students successfully reproduced 39% of the figures. The main obstacles were the need for more advanced programming skills and the incomplete documentation of the applied methods. Students were engaged, enthusiastic, and focused throughout the semester. We believe that this teaching approach will allow students to make fundamental scientific contributions under appropriate supervision. It will teach them about the scientific process, the importance of reporting standards, and the importance of openness.

Twist-open mechanism of DNA damage recognition by the Rad4/XPC nucleotide excision repair complex.

DNA damage repair starts with the recognition of damaged sites from predominantly normal DNA. In eukaryotes, diverse DNA lesions from environmental sources are recognized by the xeroderma pigmentosum C (XPC) nucleotide excision repair complex. Studies of Rad4 (radiation-sensitive 4; yeast XPC ortholog) showed that Rad4 "opens" up damaged DNA by inserting a ß-hairpin into the duplex and flipping out two damage-containing nucleotide pairs. However, this DNA lesion "opening" is slow (˜5-10 ms) compared with typical submillisecond residence times per base pair site reported for various DNA-binding proteins during 1D diffusion on DNA. To address the mystery as to how Rad4 pauses to recognize lesions during diffusional search, we examine conformational dynamics along the lesion recognition trajectory using temperature-jump spectroscopy. Besides identifying the ˜10-ms step as the rate-limiting bottleneck towards opening specific DNA site, we uncover an earlier ˜100- to 500-µs step that we assign to nonspecific deformation (unwinding/"twisting") of DNA by Rad4. The ß-hairpin is not required to unwind or to overcome the bottleneck but is essential for full nucleotide-flipping. We propose that Rad4 recognizes lesions in a step-wise "twist-open" mechanism, in which preliminary twisting represents Rad4 interconverting between search and interrogation modes. Through such conformational switches compatible with rapid diffusion on DNA, Rad4 may stall preferentially at a lesion site, offering time to open DNA. This study represents the first direct observation, to our knowledge, of dynamical DNA distortions during search/interrogation beyond base pair breathing. Submillisecond interrogation with preferential stalling at cognate sites may be common to various DNA-binding proteins.