Lysobacter enzymogenes OH11 is a non-flagellated,ubiquitous soil bacterium with broad-spectrum antifungal activities.Although lacking flagella,it employs another type of motile behavior,known as twitching motility tha...Lysobacter enzymogenes OH11 is a non-flagellated,ubiquitous soil bacterium with broad-spectrum antifungal activities.Although lacking flagella,it employs another type of motile behavior,known as twitching motility that is powered by type IV pilus(T4P)to move towards neighboring crop fungal pathogens to kill them as food.At present,little is known about how this non-flagellated bacterium controls twitching motility that is crucial for its predatory lifestyle.Herein,we present a report on how a non-canonical PilZ domain,PilZ_(Le3639),controls such motility in the non-flagellated L.enzymogenes;it failed to bind with c-di-GMP but seemed to be required for twitching motility.Using bacterial two-hybrid and pull-down approaches,we identified PilB_(Le0708),one of the PilZ_(Le3639)-binding proteins that are essential for the bacterial twitching motility,could serve as an ATPase to supply energy for T4P extension.Through site-mutagenesis approaches,we identified one essential residue of PilZ_(Le3639) that is required for its binding affinity with PilB_(Le0708) and its regulatory function.Besides,two critical residues within the ATPase catalytic domains of PilB_(Le0708) were detected to be essential for regulating twitching behavior but not involved in binding with PilZ_(Le3639).Overall,we illustrated that the PilZ-PilB complex formation is indispensable for twitching motility in a non-flagellated bacterium.展开更多
Using Arsenazo Ⅲ as a myoplasmic calcium indicator, we have studied the calcium transients evoked by voltage-clamp depolarizing pulses in frog twitch muscle fibres which had been temporarily depolarized by 80 mmol/LK...Using Arsenazo Ⅲ as a myoplasmic calcium indicator, we have studied the calcium transients evoked by voltage-clamp depolarizing pulses in frog twitch muscle fibres which had been temporarily depolarized by 80 mmol/LK^+ in the absence or presence of myoplasmic Li^+. After the high K^+ exposure, for either a short (15 rain) or long(1 h) time, the post-K^+ calcium transients could gradually be restored to the level of the pre-K^+ ones, if the fibres were not loaded with Li^+.In contrast, the post-K^+ calcium transients of Li^+-loaded fibres could not fully recover,and were depressed in a Li^+ concentration-dependent manner.The mean amplitude of the post-K^+ responses recorded more than 3.5 h after 15 min high K^+ exposure was reduced to 56% of pre-K^+ control in the fibres which had been loaded with Li^+ in 20 mmol/L Li^+ Ringer's solution.This depression could be prevented or partially reversed by exogenous myo-inositol.More depression could be induced by 1 h high K^+ exposure, but the presence of exogenous myo-inositol could not clearly prevent the post-K^+ calcium transients from reduction.Assuming that high K^+ exposure caused a depletion of myo-inositol and probably other changes in the metabolism of inositol phospholipids in Li^+ loaded fibres,we conclude that some metabolites of phosphoinositides may play modulation roles in excitation-contraction coupling in frog twitch muscle fibres.展开更多
基金supported by the Natural Science Foundation of Jiangsu Province(BK20190026,BK20181325)the National Natural Science Foundation of China(31872016)+1 种基金the Fundamental Research Funds for the Central Universities(KJJQ202001,KYT201805,KYTZ201403 and KYT202001)Jiangsu Agricultural Sciences and Technology Innovation Fund[CX(18)1003]and Innovation Team Program for Jiangsu Universities(2017).
文摘Lysobacter enzymogenes OH11 is a non-flagellated,ubiquitous soil bacterium with broad-spectrum antifungal activities.Although lacking flagella,it employs another type of motile behavior,known as twitching motility that is powered by type IV pilus(T4P)to move towards neighboring crop fungal pathogens to kill them as food.At present,little is known about how this non-flagellated bacterium controls twitching motility that is crucial for its predatory lifestyle.Herein,we present a report on how a non-canonical PilZ domain,PilZ_(Le3639),controls such motility in the non-flagellated L.enzymogenes;it failed to bind with c-di-GMP but seemed to be required for twitching motility.Using bacterial two-hybrid and pull-down approaches,we identified PilB_(Le0708),one of the PilZ_(Le3639)-binding proteins that are essential for the bacterial twitching motility,could serve as an ATPase to supply energy for T4P extension.Through site-mutagenesis approaches,we identified one essential residue of PilZ_(Le3639) that is required for its binding affinity with PilB_(Le0708) and its regulatory function.Besides,two critical residues within the ATPase catalytic domains of PilB_(Le0708) were detected to be essential for regulating twitching behavior but not involved in binding with PilZ_(Le3639).Overall,we illustrated that the PilZ-PilB complex formation is indispensable for twitching motility in a non-flagellated bacterium.
基金Project supported by the National Natural Science Foundation of China.
文摘Using Arsenazo Ⅲ as a myoplasmic calcium indicator, we have studied the calcium transients evoked by voltage-clamp depolarizing pulses in frog twitch muscle fibres which had been temporarily depolarized by 80 mmol/LK^+ in the absence or presence of myoplasmic Li^+. After the high K^+ exposure, for either a short (15 rain) or long(1 h) time, the post-K^+ calcium transients could gradually be restored to the level of the pre-K^+ ones, if the fibres were not loaded with Li^+.In contrast, the post-K^+ calcium transients of Li^+-loaded fibres could not fully recover,and were depressed in a Li^+ concentration-dependent manner.The mean amplitude of the post-K^+ responses recorded more than 3.5 h after 15 min high K^+ exposure was reduced to 56% of pre-K^+ control in the fibres which had been loaded with Li^+ in 20 mmol/L Li^+ Ringer's solution.This depression could be prevented or partially reversed by exogenous myo-inositol.More depression could be induced by 1 h high K^+ exposure, but the presence of exogenous myo-inositol could not clearly prevent the post-K^+ calcium transients from reduction.Assuming that high K^+ exposure caused a depletion of myo-inositol and probably other changes in the metabolism of inositol phospholipids in Li^+ loaded fibres,we conclude that some metabolites of phosphoinositides may play modulation roles in excitation-contraction coupling in frog twitch muscle fibres.
