Seattle Children's Attending Physician,
Immunodeficiency Clinic and Inpatient Immunology Service,
Associate Professor, Pediatrics and Adjunct Associate Professor, Immunology
Seattle Children's Research Institute
M/S C9S-7
1900 9th Avenue
Seattle, WA 98101
Tel: (206) 987-7450
E-mail Dr. Scharenberg
Dr. Scharenberg graduated summa cum laude from Indiana University with a BS in biochemistry and earned his MD with Distinction from the University of North Carolina School of Medicine. He completed postdoctoral fellowships in the Pediatric Scientist Development Program at the National Institutes of Health (NIH), the Howard Hughes Medical Institute, and in the Division of Experimental Pathology at Beth Israel Hospital, before joining the faculty of Harvard Medical School as an assistant professor.
Dr. Scharenberg is currently the co-director of the NGEC and a principal investigator in the area of Directed Evolution of Homing Endonucleases.
"The NGEC provides an opportunity to work together with a
world-class investigative team in pushing forward cutting-edge technologies for
treatment of inherited diseases of the blood."The major NGEC role of the Scharenberg lab is to receive homing endonuclease scaffolds from the Monnat and Baker labs. Scaffolds from the Monnat lab will be newly identified and characterized “wild” homing endonucleases, or novel single chain enzymes created by the Monnat lab. Scaffolds from the Baker lab will have been computationally re-designed by the Baker lab to recognize new target sites. The binding and cleavage properties of the scaffolds will be analyzed using homing endonuclease surface display technology.
By random introduction of mutations into the scaffold DNA coding sequences, the Scharenberg lab will then generate large libraries of variant homing endonucleases. These libraries will be screened using homing endonuclease surface display technology for improvements in desired properties. This process can be iterated, resulting in the gradual “evolution” of the original scaffold towards new enzymes in which the desired properties have been optimized. This process is termed “directed evolution.”
The Scharenberg lab is also working with the Rawlings lab to develop murine XSCID and IPEX animal models to be used by the Rawlings lab for its NGEC component, and with the Rawlings lab and NGEC Cell and Viral Core to develop lentiviral expression vectors for use in gene repair experiments.
The Scharenberg lab also seeks to develop platform technologies required for high throughput design and analysis of the binding and cleavage specificities of homing endonuclease.
Key lab personnel participating in this work include:
Marcia Paddock and Jordan Jarjour, grad students in the Scharenberg
lab
Approximately half of the Scharenberg lab works on directed evolution of homing endonuclease proteins, and their application to gene repair as part of the NGEC. The other half works on the role of ion channels and 2nd messengers in immune cell biology, focusing on the channel/enzyme fusion proteins TRPM2 and TRPM7. Work in the latter area is spearheaded by Jaya Sahni on TRPM7, and Yumei Song and Ben Buelow on TRPM2 and ADP-ribose, respectively.
The Scharenberg lab developed homing endonuclease surface display technology in collaboration with the Stoddard and Monnat laboratories. The lab uses surface display of homing endonucleases combined with both MACS and FACS for its work on the analysis and directed evolution of homing endonucleases.
The Scharenberg lab uses a wide variety of molecular and cellular biology methods, fluorescence imaging, flow cytometry, patch clamp, and mass spectrometry for its work on Mg2+ homeostasis and ADP-ribose 2nd messenger signaling functions.
Scharenberg AM, Humphries LA, Rawlings DJ. Calcium signalling and cell-fate choice in B cells. Nat Rev Immunol. 2007 Oct;7(10):778-89.
Sahni J, Nelson B, Scharenberg AM. SLC41A2 encodes a plasma-membrane Mg2+ transporter. Biochem J. 2007 Jan 15;401(2):505-13.
Perraud AL, Takanishi CL, Shen B, Kang S, Smith MK, Schmitz C, Knowles HM, Ferraris D, Li W, Zhang J, Stoddard BL, Scharenberg AM. Accumulation of free ADP-ribose from mitochondria mediates oxidative stress-induced gating of TRPM2 cation channels. J Biol Chem. 2005 Feb 18;280(7):6138-48. Epub 2004 Nov 23.
Schmitz C, Perraud AL, Johnson CO, Inabe K, Smith MK, Penner R, Kurosaki T, Fleig A, Scharenberg AM. Regulation of vertebrate cellular Mg2+ homeostasis by TRPM7. Cell. 2003 Jul 25;114(2):191-200.
Volna P, Jarjour J, Baxter S, Roffler SR, Monnat RJ Jr, Stoddard BL, Scharenberg AM. Flow cytometric analysis of DNA binding and cleavage by cell surface-displayed homing endonucleases. Nucleic Acids Res. 2007;35(8):2748-58. Epub 2007 Apr 10.