Elevating PI3P drives select downstream membrane trafficking pathways

Phosphoinositide signaling lipids are essential for several cellular processes. The requirement for a phosphoinositide is conventionally studied by depleting the corresponding lipid kinase. However, there are very few reports on the impact of elevating phosphoinositides. That phosphoinositides are dynamically elevated in response to stimuli suggests that, in addition to being required, phosphoinositides drive downstream pathways. To test this hypothesis, we elevated the levels of phosphatidylinositol-3-phosphate (PI3P) by generating hyperactive alleles of the yeast phosphatidylinositol 3-kinase, Vps34. We find that hyperactive Vps34 drives certain pathways, including phosphatidylinositol-3,5-bisphosphate synthesis and retrograde transport from the vacuole. This demonstrates that PI3P is rate limiting in some pathways. Interestingly, hyperactive Vps34 does not affect endosomal sorting complexes required for transport (ESCRT) function. Thus, elevating PI3P does not always increase the rate of PI3P-dependent pathways. Elevating PI3P can also delay a pathway. Elevating PI3P slowed late steps in autophagy, in part by delaying the disassembly of autophagy proteins from mature autophagosomes as well as delaying fusion of autophagosomes with the vacuole. This latter defect is likely due to a more general defect in vacuole fusion, as assessed by changes in vacuole morphology. These studies suggest that stimulus-induced elevation of phosphoinositides provides a way for these stimuli to selectively regulate downstream processes.     

A morphometric study of the carotid body in chronically hypoxic rats

We performed morphometric studies of carotid body in acutely and chronically hypoxic rats (inspired PO2 = 70 Torr, at sea level). Acute exposure was for the duration of about 10 min, and chronic exposure lasted for 28 days. We confirmed that the total volume of the organ increased by severalfold. At the light-microscopy level we found an enlargement of the volume density of the blood sinuses from 14 to 31% due to chronic hypoxia. The morphometric hematocrit increased from 39 to 70% paralleling changes in the conventionally measured venous hematocrit. These data do not show any specific plasma skimming in the carotid body blood vessels. With the electron microscope we found that the mean average volume of type I cells increased from 320 micron3 in controls to 1,120 micron3 in the chronically hypoxic rats without hyperplasia, whereas type II cells had increased in number without alteration in size. Qualitative observations revealed that the normal appearance of clusters of ovoid type I cells interspersed by capillaries had been transformed into a pattern of individual cells forming plates between expanded blood vessels with a large increase of contact area between the cells and vessels. Type II cells appeared to have proliferated without changes in individual size to cover the enlarged periphery of type I cells. The observed structural changes in the carotid body parenchyma and vasculature appear to be physiologically adaptive and provide further support for the idea that various elements in the organ are particularly sensitive to hypoxia.     

Theoretical analysis of gel electrophoretic data for interaction of lysine rich histone with supercoiled DNA.

We demonstrate the possibility of using gel electrophoresis as a technique for the quantitative analysis of interaction of lysine rich histone with DNA. On the basis of theoretical framework for extended ligand binding to one-dimensional lattices such as DNA we have set up systems of equations which relate the ligand-to-DNA ratio to the observed gel migration distance of the complex. From the analysis of experimental data for gel electrophoresis of supercoiled DNA in the presence of lysine rich histones we have found that the observed variation of electrophoretic mobilities of the histone-DNA complexes at low histone-to-DNA ratios can be described by a non-cooperative binding behaviour. At this limit we have estimated the intrinsic binding constant to be of the order of 10(3) M-1.     

DISTRIBUTION AND MULTIPLICATION OF COLONY FORMING UNITS FROM BONE MARROW AND SPLEEN AFTER INJECTION IN IRRADIATED MICE

The distribution and proliferation of CFUs from bone marrow and spleen cell suspensions were followed after injection in lethally irradiated isogeneic mice. It was found that a larger proportion of the injected bone marrow CFUs than of the spleen derived CFUs could be recovered from the recipient's spleen and femur. This consistently higher recovery points to the conclusion that a larger fraction of bone marrow-derived CFUs than of spleen-derived CFUs is capable of producing daughter CFUs, most likely due to a commitment to early differentiation of many spleen CFUs.     

SUMO fosters assembly and functionality of the MutSγ complex to facilitate meiotic crossing over

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Dopamine, sensory discharge, and stimulus interaction with CO2 and O2 in cat carotid body

It is hypothesized that carotid bodychemosensory activity is coupled to neurosecretion. The purpose of thisstudy was to examine whether there was a correspondence between carotidbody tissue dopamine (DA) levels and neuronal discharge (ND) measured from the carotid sinus nerve of perfused cat carotid bodies and tocharacterize interaction betweenCO₂ andO₂ in these responses. ND andtissue DA were measured after changing from normoxic, normocapnic control bicarbonate buffer (PO₂>120 Torr, PCO₂ 25-30 Torr, pH ~ 7.4) to normoxic hypercapnia(PCO₂ 55-57 Torr, pH7.1-7.2) or to hypoxic solutions(PO₂ 30-35 Torr) withnormocapnia (PCO₂ 25-30 Torr, pH ~ 7.4) or hypocapnia (PCO₂10-15 Torr, pH 7.6-7.8). Similar temporal changes for ND and tissue DA were found for all of the stimuli, although there was a much different proportional relationship fornormoxic hypercapnia. Both ND and DA increased above baseline valuesduring flow interruption and normocapnic hypoxia, and both decreasedbelow baseline values during hypoxic hypocapnia. In contrast, normoxichypercapnia caused an initial increase in ND, from a baseline of 175 ± 12 (SE) to a peak of 593 ± 20 impulses/s within 4.6 ± 0.9 s, followed by adaptation, whereas ND declined to 423 ± 20 impulses/s after 1 min. Tissue DA initially increased from a baselineof 17.9 ± 1.2 µM to a peak of 23.2 ± 1.2 µM within 3.0 ± 0.7 s, then declined to 2.6 ± 1.0 µM. The substantialdecrease in tissue DA during normoxic hypercapnia was not consistentwith the parallel changes in DA with ND that were observed for hypoxic stimuli.