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• 1. [Article] Non-canonical Binding of Calmodulin to Aquaporin-0: Implications for Channel Regulation

  Aquaporins (AQPs) are a family of ubiquitous membrane channels that conduct water across cell membranes. AQPs form homo-tetramers containing four functional and independent water pores. Aquaporin-0 (AQP0) ...

  Citation × Citation Citation Abstract Copy Title: Non-canonical Binding of Calmodulin to Aquaporin-0: Implications for Channel Regulation Author: Reichow, Steve, Gonen, Tamir Year: 2008 Aquaporins (AQPs) are a family of ubiquitous membrane channels that conduct water across cell membranes. AQPs form homo-tetramers containing four functional and independent water pores. Aquaporin-0 (AQP0) is expressed in the eye lens where its water permeability is regulated by calmodulin (CaM). Here we use a combination of biochemical methods and NMR spectroscopy to probe the interaction between AQP0 and CaM. We show CaM binds the AQP0 C-terminal domain in a calcium dependent manner. We demonstrate that only two CaM molecules bind a single AQP0 tetramer in a non-canonical fashion, suggesting a form of co-operativity between AQP0 monomers. Based on these results, we derive a structural model of the AQP0/CaM complex, which suggests CaM may be inhibitory to channel permeability by capping the vestibules of two monomers within the AQP0 tetramer. Finally, phosphorylation within AQP0’s CaM binding domain inhibits the AQP0/ CaM interaction suggesting a temporal regulatory mechanism for complex formation. Title: Non-canonical Binding of Calmodulin to Aquaporin-0: Implications for Channel Regulation Author: Reichow, Steve, Gonen, Tamir Year: 2008 Aquaporins (AQPs) are a family of ubiquitous membrane channels that conduct water across cell membranes. AQPs form homo-tetramers containing four functional and independent water pores. Aquaporin-0 (AQP0) is expressed in the eye lens where its water permeability is regulated by calmodulin (CaM). Here we use a combination of biochemical methods and NMR spectroscopy to probe the interaction between AQP0 and CaM. We show CaM binds the AQP0 C-terminal domain in a calcium dependent manner. We demonstrate that only two CaM molecules bind a single AQP0 tetramer in a non-canonical fashion, suggesting a form of co-operativity between AQP0 monomers. Based on these results, we derive a structural model of the AQP0/CaM complex, which suggests CaM may be inhibitory to channel permeability by capping the vestibules of two monomers within the AQP0 tetramer. Finally, phosphorylation within AQP0’s CaM binding domain inhibits the AQP0/ CaM interaction suggesting a temporal regulatory mechanism for complex formation. Close

• 2. [Article] Cooperative Interaction of Transcription Termination Factors with the RNA Polymerase II C-terminal Domain

  Phosphorylation of the C-terminal domain of RNA polymerase II controls the co-transcriptional assembly of RNA processing and transcription factors. Recruitment relies on conserved CTDinteracting domains ...

  Citation × Citation Citation Abstract Copy Title: Cooperative Interaction of Transcription Termination Factors with the RNA Polymerase II C-terminal Domain Author: Lunde, Bradley M., Reichow, Steve, Kim, Minkyu, Suh, Hyunsuk, Leeper, Thomas C., Yang, Fan, Mutschler, Hannes, Buratowski, Stephen, Meinhart, Anton, Varani, Gabriele Year: 2010 Phosphorylation of the C-terminal domain of RNA polymerase II controls the co-transcriptional assembly of RNA processing and transcription factors. Recruitment relies on conserved CTDinteracting domains that recognize different CTD phosphoisoforms during the transcription cycle, but the molecular basis for their specificity remains unclear. We show that the CTD-interacting domains of two transcription termination factors, Rtt103 and Pcf11, achieve high affinity and specificity both by specifically recognizing the phosphorylated CTD and by cooperatively binding to neighboring CTD repeats. Single amino acid mutations at the protein-protein interface abolish cooperativity and affect recruitment at the 3′-end processing site in vivo. We suggest that this cooperativity provides a signal-response mechanism to ensure that its action is confined only to proper polyadenylation sites where Serine 2 phosphorylation density is highest. Title: Cooperative Interaction of Transcription Termination Factors with the RNA Polymerase II C-terminal Domain Author: Lunde, Bradley M., Reichow, Steve, Kim, Minkyu, Suh, Hyunsuk, Leeper, Thomas C., Yang, Fan, Mutschler, Hannes, Buratowski, Stephen, Meinhart, Anton, Varani, Gabriele Year: 2010 Phosphorylation of the C-terminal domain of RNA polymerase II controls the co-transcriptional assembly of RNA processing and transcription factors. Recruitment relies on conserved CTDinteracting domains that recognize different CTD phosphoisoforms during the transcription cycle, but the molecular basis for their specificity remains unclear. We show that the CTD-interacting domains of two transcription termination factors, Rtt103 and Pcf11, achieve high affinity and specificity both by specifically recognizing the phosphorylated CTD and by cooperatively binding to neighboring CTD repeats. Single amino acid mutations at the protein-protein interface abolish cooperativity and affect recruitment at the 3′-end processing site in vivo. We suggest that this cooperativity provides a signal-response mechanism to ensure that its action is confined only to proper polyadenylation sites where Serine 2 phosphorylation density is highest. Close