TY  - JOUR
A1  - Panzer, Sabine
A1  - Zhang, Chong
A1  - Konte, Tilen
A1  - Bräuer, Celine
A1  - Diemar, Anne
A1  - Yogendran, Parathy
A1  - Yu-Strzelczyk, Jing
A1  - Nagel, Georg
A1  - Gao, Shiqiang
A1  - Terpitz, Ulrich
T1  - Modified Rhodopsins From Aureobasidium pullulans Excel With Very High Proton-Transport Rates
JF  - Frontiers in Molecular Biosciences
N2  - Aureobasidium pullulans is a black fungus that can adapt to various stressful conditions like hypersaline, acidic, and alkaline environments. The genome of A. pullulans exhibits three genes coding for putative opsins ApOps1, ApOps2, and ApOps3. We heterologously expressed these genes in mammalian cells and Xenopus oocytes. Localization in the plasma membrane was greatly improved by introducing additional membrane trafficking signals at the N-terminus and the C-terminus. In patch-clamp and two-electrode-voltage clamp experiments, all three proteins showed proton pump activity with maximal activity in green light. Among them, ApOps2 exhibited the most pronounced proton pump activity with current amplitudes occasionally extending 10 pA/pF at 0 mV. Proton pump activity was further supported in the presence of extracellular weak organic acids. Furthermore, we used site-directed mutagenesis to reshape protein functions and thereby implemented light-gated proton channels. We discuss the difference to other well-known proton pumps and the potential of these rhodopsins for optogenetic applications.
KW  - black yeast
KW  - photoreceptor
KW  - microbial rhodopsins
KW  - optogenetics
KW  - proton channel
KW  - membrane trafficking
KW  - fungal rhodopsins
KW  - Aureobasidium
Y1  - 2021
U6  - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-249248
SN  - 2296-889X
VL  - 8
ER  - 
TY  - JOUR
A1  - Zhou, Yang
A1  - Ding, Meiqi
A1  - Duan, Xiaodong
A1  - Konrad, Kai R.
A1  - Nagel, Georg
A1  - Gao, Shiqiang
T1  - Extending the Anion Channelrhodopsin-Based Toolbox for Plant Optogenetics
JF  - Membranes
N2  - Optogenetics was developed in the field of neuroscience and is most commonly using light-sensitive rhodopsins to control the neural activities. Lately, we have expanded this technique into plant science by co-expression of a chloroplast-targeted β-carotene dioxygenase and an improved anion channelrhodopsin GtACR1 from the green alga Guillardia theta. The growth of Nicotiana tabacum pollen tube can then be manipulated by localized green light illumination. To extend the application of analogous optogenetic tools in the pollen tube system, we engineered another two ACRs, GtACR2, and ZipACR, which have different action spectra, light sensitivity and kinetic features, and characterized them in Xenopus laevis oocytes, Nicotiana benthamiana leaves and N. tabacum pollen tubes. We found that the similar molecular engineering method used to improve GtACR1 also enhanced GtACR2 and ZipACR performance in Xenopus laevis oocytes. The ZipACR1 performed in N. benthamiana mesophyll cells and N. tabacum pollen tubes with faster kinetics and reduced light sensitivity, allowing for optogenetic control of anion fluxes with better temporal resolution. The reduced light sensitivity would potentially facilitate future application in plants, grown under low ambient white light, combined with an optogenetic manipulation triggered by stronger green light.
KW  - optogenetics
KW  - rhodopsin
KW  - light-sensitive anion channel
KW  - surface potential recording
KW  - pollen tube
Y1  - 2021
U6  - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-236617
SN  - 2077-0375
VL  - 11
IS  - 4
ER  -