Protein Quality Control:

How to Recognize and Remove Misfolded and Misassembled Proteins

   Before a protein can play its destined part in a cell, it must successfully complete a few fundamental processes: correct folding into its destined three-dimensional structure, integration into its larger protein complex (where applicable) and completion of any additional chemical alterations. Such posttranslational modifications entail for example the addition of disulfide, glycans or lipids. But what happens if something during these processes goes wrong? What if a protein is not properly translated, folded, assembled into complexes, or chemically modified? The resulting dysfunctional proteins, if left unchecked, pose a serious threat to a cell’s health and ultimately life. It is thus of paramount importance for the cell to control and maintain the quality of its protein production. In fact, many human diseases are linked to problems in this cellular quality control process that impede proper removal of dysfunctional proteins or permit their accumulation and aggregation.

   The long-term aim of our group’s research is to understand the processes that control and maintain the quality of proteins produced in the endoplasmic reticulum (ER). The cell’s ER is responsible for the proper folding and chemical modification of a third of all eukaryotic proteins including all membrane-embedded and secreted proteins. Also, certain posttranslational modifications such as disulfide or glycans addition are exclusive to the ER. But what happens to all the dysfunctional proteins that are unavoidably produced there as well? They are processed by the ER-associated degradation (ERAD) pathway. Embedded in the ER membrane, the multi-protein ERAD complex is tasked with identifying, unfolding, and transporting corrupt proteins into the cytoplasm, where they are passed on to the machinery of the proteasome.

Our lab’s research into the cellular management of protein fidelity focuses on the identification, unfolding, and translocation of dysfunctional proteins by the ERAD. We want to understand how these key steps are regulated and controlled because it provides a potential avenue to precisely modify the ERAD’s response instead of just terminating it. This marks the proteins involved in these steps as the ideal targets to selectively manipulate protein degradation. Such therapeutic approaches have great potential for diseases where for example protein overproduction -like in cancer- is a key issue or where specific proteins form detrimental aggregates as in certain forms of amyotrophic lateral sclerosis (ALS).  

In our current projects, we focus specifically on the important ERAD proteins derlin, selenoprotein K (SELENOK) and selenoprotein S (SELENOS, VIMP), their interactions with each other as well as other proteins and how their actions and interactions contribute to ERAD functions and their control. The specific research directions are described in more detail below.

Derlin: The Essential, yet Mysterious ERAD Component 

  The very core of the ERAD complex is formed by ubiquitin ligases (enzymes that tag ubiquitin to proteins) and the membrane protein derlin. Like all members of the family of rhomboid proteins, derlin can both recognize proteins inside the lipid bilayers and locally alter the properties of the membrane. Because of these key features, rhomboid proteins are often involved in the regulation of membrane proteins. The specifics of these regulatory processes, 

however, are quite different between family members and can involve for example the cleaving of proteins, releasing of protein fragments from the membrane, controlling trafficking, or -like derlin- deforming proteins for their ultimate regulation. When it comes to derlin, we know that it is essential for the degradation of some classes of ERAD substrates but not all. Originally, it was assumed that derlin’s primary function was to recognize misfolded membrane proteins and target them for degradation. Recently, however, it has become questionable that this is indeed its only or even primary function. An alternative hypothes regarding the role of derlin in the ERAD asserts that derlin together with several additional proteins might be forming a path for moving substrates across the ER membrane. Still others propose that it can operate indeed independently to translocate proteins across the membrane under yet unknown conditions or maybe in the case of particular substrates.

To ultimately establish derlin’s overall function in the ERAD our group focuses on answering a series of interconnected questions:
      • How does derlin -either by itself or together with its ancillary proteins- recognize misfolded membrane proteins?
      • Which of derlin’s protein partners are responsible for controlling the specificity and/or stability of the recognition process?
      • How does derlin ‘deform’ a recognized protein and enable it to move out of the membrane?
      • How does derlin modulate the properties of its surrounding membrane?
      • How are all of these functions connected to derlin’s structure?

Selenoprotein K : A Relay Station for Pathways at the ER Membrane

   Selenoprotein K (SELENOK) is a small intrinsically disordered signaling protein. It is involved both in protein quality control but also in the attachment of hydrophobic chains to proteins which targets them to membranes. Because selenoprotein K has the rare amino acid selenocysteine, it is most likely an enzyme. Like other proteins we study, it is embedded in the ER membrane. Unpublished results from our group show that selenoprotein K catalyzes the cleavage of its own peptide bond. Interestingly,the cleavage releases the selenocysteine-containing segment which could terminate the proteins enzymatic activity. The fact that the same selenocysteine-containing segment is also cleaved by a different cellular protease during signal transduction suggest a regulatory function of selenoprotein K’s autoproteolytic activity.

Building on our discovery our lab is now working on several related questions regarding the chemistry and biological roles of this autocleavage:

• Why does the segment with selenocysteine (and the enzymatic activity) need to be regulated in such a multi-faceted process?
• What happens to the cleaved segments in the cell? Is it carrying a specific function or is it merely degraded?
• Can protein partners of SELENOK change the rate or location of the cleavage sites?

Selenoprotein S: A Signaling Hub at the Heart of ERAD Initiation

   The integral membrane Selenoprotein S ( SELENOS, VIMP) can turn particular functions of the ERAD on and off. Selenoprotein S does so by recruiting the ATPase p97 to the complex and thereby providing the energy source to unfold and move misfolded proteins from the ER into the cytoplasm.It may also recruit proteins that assist in processing dysfunctional proteins. Furthermore we know that in vitro it can act as an enzyme however its enzymatic activity in the cell remains still obscure. Selenoprotein S plays a role in overall human health becasue it is an essential gene and mutations of this proteins significantly raise the risk for cardiovascular diseases although the details of    

this connection remain currently still elusive. From a structural perspective SELENOS is interesting because it contains both ordered and disordered segments as well as membrane-bound regions. In our group we work to find answers to the following questions:

      • What proteins are recruited by the different SELENOS segments and what functions do these proteins coordinate?
      • Why does SELENOS occur in two forms: one with and one without selenocysteine?
      • What is the enzymatic function of SELENOS in vivo?
      • Can inhibition of SELENOS enzymatic function be used to block selected ERAD functions. Does this leads to significantly                        increased stress in fast multiplying cancer cells and ultimately to new therapeutic approaches?