Biology subjects

Jiang, Q.-X.

Publications and source records attributed to Jiang, Q.-X..

4 recordsLinked to original sources

bioRxiv2024

Molecular requirements of chromogranin B for the long-sought anion shunter of regulated secretion

All eukaryotes utilize regulated secretion to release molecular signals packaged in secretory granules for local and remote signaling. An anion shunt conductance was first suggested in secretory granules of bovine chromaffin cells nearly five decades ago. Biochemical identity of this conductance remains undefined. CLC-3, an intracellular Cl-/H+ exchanger, was proposed as a candidate sixteen years ago, which, however, was contested experimentally. Here, we show that chromogranin B (CHGB) makes the kernel of the long-sought anion shunter in cultured and primary neuroendocrine cells and its channel functions are essential to proper granule maturation. Intragranular pH measurements and cargo maturation assays revealed that normal granular acidification, proinsulin-insulin conversion, and dopamine-loading in neuroendocrine cells all rely on functional CHGB+ channels. Primary {beta}-cells from Chgb-/- mice exhibited persistent granule deacidification, which suffices to uplift plasma proinsulin level, diminish glucose-induced 2nd-phase insulin secretion and dwindle monoamine content in chromaffin granules from the knockout mice. Data from targeted genetic manipulations, dominant negativity of a deletion mutant lacking channel-forming parts and tests of CLC-3/5 and ANO-1/2 all exclude CHGB-less channels from anion shunting in secretory granules. The highly conserved CHGB+ channels thus function in regulated secretory pathways in neuronal, endocrine, exocrine and stem cells of probably all vertebrates. HIGHLIGHTSO_LILoss of CHGB channel functions impairs secretory granule acidification in neuroendocrine cells, which necessitates anion shunt conduction. C_LIO_LICHGB{Delta}MIF, a mutant unable to form a functional Cl- channel, exerts negative dominance on endogenous CHGB and results in granule deacidification in cultured cells. C_LIO_LINeither CLC-3 & -5 nor ANO-1 & -2 participate in the CHGB-mediated granule acidification. Clcn3 knockout effects on regulated secretion can be attributed to its functions in endosomal and endolysosomal compartments. C_LIO_LIPrimary Chgb-/- {beta}-cells exhibit persistent granule deacidification, presenting a unifying mechanism for disparate mouse phenotypes: hyperproinsulinemia, near abrogation of 2nd phase insulin release after glucose challenge and diminution of monoamine contents in chromaffin granules. C_LI

cell biology↗

bioRxiv2019

Membrane insertion of chromogranin B for granule maturation in regulated secretion

Regulated secretion serves responses to specific stimuli in eukaryotes. An anion conductance was found essential for maturation and acidification of secretory granules four decades ago, but its genetic identity was unknown. We now demonstrate that chromogranin B (CHGB), an obligate granule protein, constitutes the long-sought anion channel. High-pressure freezing immuno-electron microscopy and biochemical assays showed native CHGB in close proximity to secretory granule membranes, and its membrane-bound and soluble forms both reconstituted Cl- channels. Release of secretory granules delivered CHGB clusters to plasma membranes, which dominate whole-cell anion conductance. Intragranular pH measurements and cargo maturation assays found that CHGB channels supported proinsulin - insulin conversion and dopamine-loading in neuroendocrine cells. {beta}-cells from Chgb-/- mice exhibited significant granule deacidification, accounting for hyperproinsulinemia, altered glucose-tolerance response and lower dopamine concentration in chromaffin granules in these animals. Membrane insertion of well-conserved CHGB is thus indispensable for granule maturation in exocrine, endocrine and neuronal cells. HighlightsO_LINative CHGB is amphipathic and distributes in the lumen and membranes of secretory granules with contrastingly different destinies and functions. C_LIO_LINative CHGB, once delivered to cell surface via granule exocytosis, dominates anion conductance in plasma membranes. C_LIO_LICHGB channels facilitate granule acidification and cargo maturation in cultured and primary neuroendocrine cells. C_LIO_LICHGB channels from bovine, rat and mouse cells all serve the long-missing, intra-organellar anion shunt pathway in the secretory granules for regulated secretion. C_LI

cell biology↗

bioRxiv2019

A dynamic control of human telomerase holoenzyme

Human telomerase functions in maintaining genome stability by adding telomeric repeats to the termini of linear chromosomes. Past studies have revealed profound insights into telomerase functions. However, low abundance of functional telomerase and difficulty in quantifying its activity leave partially characterized its thermodynamic and kinetic properties. Using a newly developed method to count individual extension products, we demonstrate that human telomerase holoenzymes contain fast- and slow-acting catalytic sites. Surprisingly, both active sites become inactive after two consecutive rounds of catalysis. The fast active sites turn off ~40-fold quicker than the slow ones and exhibit higher affinity to substrates. In dimeric enzymes, the two sites work in tandem with the faster site functioning before the slower one. In monomeric enzymes, the active sites also perform single-run catalysis. Interestingly, the inactive enzymes can be reactivated by intracellular telomerase-activating factors (iTAFs) available in multiple cell types. Together, the single-run catalysis and the iTAF-triggered reactivation serve as a novel control circuit to ensure that the telomerase holoenzymes are dynamically controlled to match their number of active sites with the number of telomeres they extend. Such exquisite kinetic control of telomerase activity is expected to play important roles in cell division and ageing.

cell biology↗

bioRxiv2018

Novel organelle anion channels formed by chromogranin B drive normal granule maturation in endocrine cells

All endocrine cells need an anion conductance for maturation of secretory granules. Identity of this family of anion channels has been elusive for forty years. We now show that a family of granule protein, CHGB, serves the long-sought conductance. CHGB interacts with membranes through two amphipathic helices, and forms a chloride channel with large conductance and high anion selectivity. Fast kinetics and high cooperativity suggest that CHGB tetramerizes to form a functional channel. Nonfunctional mutants separate CHGBs function in granule maturation from that in granule biogenesis. In neuroendocrine cells, CHGB channel and a H+-ATPase drives normal insulin maturation inside or dopamine loading into secretory granules. CHGBs tight membrane-association after exocytotic release of secretory granules separates its intracellular function from extracellular functions of its proteolytic peptides. CHGB-null mice show consistent impairment of granule acidification in pancreatic beta-cells. These findings together support that the phylogenetically conserved CHGB proteins constitute a new family of organelle chloride channels in the regulated secretory pathway among various endocrine cells.

cell biology↗