Here we report that Mdm38 and Mba1 display overlapping functions in mitochondrial protein expression. Both Mdm38 and Mba1 interact with mitochondrial ribosomes and are required for translation of COX1 and cytochrome b mRNAs.

The COVID‐19 pandemic changed health‐care operations around the world and has interrupted standard clinical practices as well as created clinical research challenges for cancer patients. Cancer patients are uniquely susceptible to COVID‐19 infection and have some of the worst outcomes. Importantly, cancer therapeutics could potentially render cancer patients more susceptible to demise from COVID‐19 yet the poor survival outcome of many cancer diagnoses outweighs this risk. In addition, the pandemic has resulted in risks to health‐care workers and research staff driving important change in clinical research operations and procedures. Remote telephone and video visits, remote monitoring, electronic capture of signatures and data, and limiting sample collections have allowed the leadership in our institution to ensure the safety of our staff and patients while continuing critical clinical research operations. Here we discuss some of these unique challenges and our response to change that was necessary to continue cancer clinical research; and, the impacts the pandemic has caused including increases in efficiency for our cancer research office.

and Peter Rehling, peter.rehling@medizin.uni-goettingen.de † These authors contributed equally to this work.Mitochondrial ribosomes synthesize core subunits of the inner membrane respiratory chain complexes. In mitochondria, translation is regulated by mRNA-specific activator proteins and occurs on membrane-associated ribosomes. Mdm38/Letm1 is a conserved membrane receptor for mitochondrial ribosomes and specifically involved in respiratory chain biogenesis. In addition, Mdm38 and its higher eukaryotic homolog Letm1, function as K + /H + or Ca 2+ /H + antiporters in the inner membrane. Here, we identify the conserved ribosome-binding domain (RBD) of Mdm38 and determine the crystal structure at 2.1Å resolution. Surprisingly, Mdm38 RBD displays a 14-3-3-like fold despite any similarity to 14-3-3-proteins at the primary sequence level and thus represents the first 14-3-3-like protein in mitochondria. The 14-3-3-like domain is critical for respiratory chain assembly through regulation of Cox1 and Cytb translation. We show that this function can be spatially separated from the ion transport activity of the membrane integrated portion of Mdm38. On the basis of the phenotypes observed for mdm38 as compared to Mdm38 lacking the RBD, we suggest a model that combining ion transport and translational regulation into one molecule allows for direct coupling of ion flux across the inner membrane, and serves as a signal for the translation of mitochondrial membrane proteins via its direct association with the protein synthesis machinery.

I herewith declare that this thesis has been written independently and with no other sources and aids than explicitly quoted.Christine Vollmer 11 1.2.2 Mitochondrial ribosomes 13 1.2.3 Mitochondrial protein export machinery 15 1.2.4 Mitochondrial translation 18 1.2.4.1 Regulation of mitochondrial translation 21 1.2.5 Mdm38 24 1.3 Aims of the work 27 2. MATERIAL & METHODS 28 2.1 Material 28 2.1.1 Chemicals, reagents and enzymes 28 2.1.2 Disposables 31 2.1.3 Kits 32 2.1.4 Laboratory equipment 32 2.1.5 Vectors 34 2.1.6 Antibodies 34 2.1.7 Microorganisms 36 2.1.7.1 E. coli strains 36 2.1.7.2 S. cerevisiae strains 36 2.2 Media & growth conditions 37 2.2.1 Media and growth conditions for E. coli 37 2.2.1.1 Media for E. coli 37 2.2.1.2 Cultivation of E. coli 40 2.2.1.3 Growth phase analysis of cultures 41 2.2.1.4 Preparation of permanent (glycerol) cryo--stocks 41 2.2.2 Media and growth conditions for S. cerevisiae 41 2.2.2.1 Media for S. cerevisiae 41 2.2.2.2 Cultivation of S. cerevisiae 42 2.2.2.3 Growth test of S. cerevisiae on agar plates 42 2.2.2.4 Preparation of cryo--stocks 43 2.2.2.5 Isolation of mitochondria from S. cerevisiae 43 2.3 Methods in molecular biology 44 2.3.1 Isolation of DNA 44 2.3.1.1 Isolation of yeast genomic DNA 44 2.3.1.2 Isolation of plasmid DNA from E. coli 45 2.3.1.3 Measurement of DNA concentration 45 2.3.2 Cloning of DNA fragments 45 2.3.2.1 DNA amplification by Polymerase Chain Reaction (PCR) 4.1. Function of Mdm38-Mba1 interaction within the mitochondrial translation machinery 105 4.2 Mdm38: The first putative mitochondrial 14-3-3-like protein in S. cerevisiae 110 4.3 Implications between yeast and human proteins Mdm38 and LETM1 116 4.4 Functional model of Mdm38 119 5. SUMMARY 122 REFERENCES 123 ABBREVATIONS 142 CURRICULUM VITAE 145 LIST OF TABLES 1.1 Overview of the five best--studied mRNA translation activator interactions in the yeast S. cerevisiae 19 2.1 Most often used primary antibodies 35 2.2 Auto--induction medium 38 2.3 Reaction setups for FideliTaq and KOD polymerases 46 2.4 PCR--program for FideliTaq and KOD polymerases 46 2.5 PCR--program for sequencing of DNA 49 2.6 Cloning of Mdm38 and LETM1 constructs 50 2.7 Primer sequences used for generation of truncation constructs 50 2.8 Components for 1.5 x translation buffer 63 3.1Data collection and refinement statistics of the Mdm38 CTD crystal 84 Mitochondrial protein translocasesAs mitochondria are involved in numerous cellular processes (see 1.1.2), a huge number of proteins are required to fulfill all their tasks. Recent studies revealed that the