Shashiva7 Phosphoinositides

nibbs06's version from 2016-04-16 22:51


Question Answer
What are phosphoinositides key players in regulating cellular processes? Cellular proliferation, Cell death, Cell motility, Intracellular membrane trafficking internalization - endocytosis (lysosomes) & Cellular signaling and metabolism.
Where do phosphoinositides originate from? (mother molecule) Phosphatidylinositol (fatty acid residues)
Where OH present in 3,4,5 positions tells us? Where it can be phosphorylated (PI 3-K, PI 4-K, PI 5-K)
What family contains monophosphoinosities? Phosphoinositides 3P, 4P, 5P
What family contains bisphosphoinositides? Phosphoinositides 3,4P2, 4,5P2, 3,5P2
What family contains trisphosphoinositides? Phosphoinositides 3,4,5P3
What is the difference between phosphatidylinositol (mother molecule) vs phosphoinositides (children? Phosphatidylinositol does not have phosphates
True or False. Phosphoinositides have a low abundance in cells – roughly 1% of total phospholipids.True
How can we detect phosphoinositide abundance? Best – quantify – HPLC – high pressure lipid chromatography. Must first label cells (P52 that will go to phosphorylated molecule)
What are the 2 most abundant phosphoinositides? Phosphoinositide 4-P and phosphoinositide 4,5-P2
How may we identify phosphoinositide 5-P? Use PtdIns 5-P mass assay
What is another way of getting good resolution for PtdIns5-P quantification? Place it in vitro, add a kinase (type 2 PIK) to form the PtdIns 4,5-P2 and then resolve by TLC and confirm by HPLC
What is a form of direct detection to quantify? Mass spectrometry (MS)
Why is it hard to find levels of phosphoinositides? The kinases and phosphatases are interconvertable and go through constant turnover – they are not static and have arrows in all directions
What makes phosphoinositides versatile? They are amphoteric, Membrane bound, Low abundant molecules, Communicate via second messengers & Localized – transmit further messaging
How can we detect phosphoinositide localization in intact cells? Antibodies – detected by immunofluorescence using a fluorescent tag. Bio-reporters for specific interactions – tagged with fluorescent marker & Indirectly by enzyme localization.
Which molecules do not have bioreporters to accurately detect their localization? PtdIns 5P and PtdIns 3,5P2 (their localization is presumed)
What is PIP3 known as? Where is it located? PI(3,4,5)P3 & Plasma membrane
What is the precursor of PI(3,5)P2? PI3P
What does GFP with fluorescent microscopy show us? Location of the phosphoinositides, P(4,5)P2 – on plasma membrane, P(4)P – on transgolgi, P(3)P – sporadic lighting throughout (endosome)
Which binding molecules contain 2% of human genes? FYVE, PX
True or False. PI’s are asymmetrically localized on membranes.True
What is the main PI that generates PIP3? PI(4,5)P2
What are the PI’s that are expressed in the endosome stage? PI3P, PI3,5P2, PI4,5P2, PI5P, PI4P, PI3,4P2
What are the mammalian PI kinases? Phosphoinositide-3-kinases, Phosphoinositol 4-kinases, Phosphoinositide 5-kinase, PIKFYVE – binds PI3P (have domains and invariant kinases) – puts a phosphate in the 5 position
What is the spliced forms vs isoforms difference? Spliced – come from one gene
Isoform – alternative genes? Shashiva discovered SAC3, which dephosphorylates to generate PI3