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Molecular genetics group (Head: K.P. Knobeloch)

Our group aims to understand the molecular and biological function of distinct components of the Ubiquitin- and Ubiquitin like systems within the context of the whole organism. Therefore we generate and analyze conditional Knockout and Knock-In models with a special focus on the ISG15 modification system and Ubiquitin specific proteases.

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Research Topics

Interferon stimulated gene 15 (ISG15)

It is well established that posttranslational modification by ubiquitin serves as a basic mechanism to control a wide range of cellular functions. Analogous to ubiquitin also other proteins with structural similarity- so called Ubiquitin like proteins (UBL) - can be covalently attached to target proteins and modify their function. Examples are SUMO, NEDD8 or FAT10. Interferon-stimulated gene 15 (ISG15/UCRP) was the first UBL described and represents one of the genes most strongly upregulated upon IFN induction. ISG15 is conjugated to a wide variety of target proteins and “Isgylation” is mediated by the activity of E1, E2 and E3 ligases. The conjugation represents a reversible process and ubiquitin specific protease 18 (USP18/UBP43) - which was originally described as an ubiquitin deconjugating enzyme - was reported to be an ISG15 specific isopeptidase. Using knockout animals generated in our lab, we investigate the function of the ISG15 modification system. We  were able to show that while ISG15 is dispensable for antiviral activity against VSV and LCMV it serves as a critical molecule in the defence against Influenza and Herpes infections. The functional role of the interplay between ISG  modification and the ubiquitin system on the ligase and deconjugation level is currently under investigation.

References:
Lenschow DJ, Lai C, Frias-Staheli N, Giannakopoulos NV, Lutz A, Wolff T, Osiak A, Levine B, Schmidt RE, García-Sastre A, Leib DA, Pekosz A, Knobeloch KP, Horak I, Virgin HW 4th. IFN-stimulated gene 15 functions as a critical antiviral molecule against influenza, herpes, and Sindbis viruses. Proc Natl Acad Sci U S A. 2007 Jan 23;104(4):1371-6.

Knobeloch KP, Utermohlen O, Kisser A, Prinz M, Horak I. Reexamination of the role of ubiquitin-like modifier ISG15 in the phenotype of UBP43-deficient mice. Mol Cell Biol. 2005 Dec;25(24):11030-4.

Osiak A, Utermohlen O, Niendorf S, Horak I, Knobeloch KP. ISG15, an interferon-stimulated ubiquitin-like protein, is not essential for STAT1 signaling and responses against vesicular stomatitis and lymphocytic choriomeningitis virus. Mol Cell Biol. 2005 Aug;25(15):6338-45.

Ubiquitin specific poteases (USPs)

In analogy to phosphatases in protein phosphorylation ubiquitination of proteins is counteracted by the activity of Ubiquitin deconjugating enzymes (DUBs). Although much progress has been made in characterizing enzymes that link ubiquitin to proteins, the understanding of deubiquitinating enzymes is just beginning to evolve. The human genome contains more than 80 different DUBs, most of them belonging to the family of Ubiquitin specific proteases (USPs).

Figure: Editing functions of deubiquitinating enzymes. Deubiquitinating enzymes may negatively regulate proteolysis or other signaling functions of ubiquitination such as internalization or altered protein function by removing the ubiquitin chain from the target proteins.

The Ubiquitin isopeptidase UBPY (USP8) represents a particular interesting member of deubiquitinating enzymes as the molecule is growth regulated and contains a structural motif for SH3 domain binding. Using conditional mutagenesis we generated mice that allow the time and cell specific inactivation of UBPY in the context of the whole organism. We were able to show that lack of UBPy results in embryonic lethality, whereas its induced inactivation in adults causes fatal liver failure. The defect is accompanied by a strong reduction or absence of several growth factor receptor tyrosine kinases (RTKs) like epidermal growth factor receptor (EGFR), hepatocyte growth factor receptor (c-met) and ERBB3. Consequently embryonic fibroblasts undergo growth arrest upon induced deletion of UBPy. UBPy deficient cells exhibit aberrantly enlarged early endosomes colocalizing with enhanced ubiquitination and have reduced levels of HRS and STAM2. Collectively our results demonstrate that UBPy is essential for receptor tyrosine kinase stability and to maintain proper endosomal transport in vivo. Currently these mice are used to elucidate the function of UBPY specifically in the brain and diverse subsets of immune relevant cell types.

Reference:
Niendorf S, Oksche A, Kisser A, Löhler J, Prinz M, Schorle H, Feller S, Lewitzky M, Horak I, Knobeloch KP. Essential role of ubiquitin-specific protease 8 for receptor tyrosine kinase stability and endocytic trafficking in vivo. Mol Cell Biol. 2007 Jul;27(13):5029-39.

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