Cell structure in the context of insulin secretion Flashcards
Glucose-Induced Insulin Secretion
1️⃣ Glucose Uptake
Glucose enters β-cells via GLUT2 transporter.
2️⃣ Glucose Metabolism
Glucose is metabolized to produce ATP, increasing the ATP:ADP ratio.
3️⃣ ATP-Dependent K⁺ Channel Closure
Increased ATP levels cause KATP channels to close, leading to cell depolarization.
4️⃣ Membrane Depolarization
The closure of KATP channels leads to plasma membrane depolarization.
5️⃣ Calcium Influx
Depolarization opens voltage-dependent Ca²⁺ channels (VDCC), allowing Ca²⁺ influx.
6️⃣ Increase in Intracellular Ca²⁺
The rise in Ca²⁺ triggers insulin granule movement.
7️⃣ Insulin Secretion
Insulin-containing granules fuse with the plasma membrane, releasing insulin into the bloodstream.
Insulin Secretory Granule Biogenesis
1️⃣ Gene Transcription
INS gene is transcribed into preproinsulin mRNA.
2️⃣ Translation in Rough ER
Preproinsulin is synthesized in the rough endoplasmic reticulum (ER).
3️⃣ Processing in ER
Preproinsulin is cleaved to form proinsulin.
4️⃣ Transport to Golgi
Proinsulin is transported to the trans-Golgi network.
5️⃣ Secretory Granule Formation
Proinsulin is processed into insulin and C-peptide in immature granules.
6️⃣ Maturation of Secretory Granules
Insulin forms hexamers with Zn²⁺, leading to mature granule formation.
7️⃣ Storage & Release
Mature insulin granules are stored and later released upon glucose stimulation.
How are Insulin SGs Prioritised for Secretion?
1. Proximity to the plasma membrane, docking.
Insulin granules exist in different pools based on their location and readiness for release.
Reserve Pool (~11,600 granules):
Deep inside the cell, these granules are not immediately available for secretion.
Morphologically Docked Pool (~1,300 granules):
Granules that have moved closer to the plasma membrane but are not fully ready for release.
Readily Releasable Pool (RRP, ~100 granules):
Already docked at the membrane and can be rapidly released when stimulated by Ca²⁺ influx.
Process:
Some granules are already docked and “primed” for exocytosis.
When glucose increases, Ca²⁺ triggers exocytosis of these docked granules.
Other granules from deeper pools move forward to replenish the docked pool.
only a small fraction of SG are ready for exocytosis …
The rest are either:
1️⃣ Not yet docked at the membrane (so they need time to move forward).
2️⃣ Not fully matured or primed for release (some granules need additional signaling to be activated).
3️⃣ Being held in reserve to ensure insulin is available for future demands.
How are Insulin SGs Prioritised for Secretion?
Restless Newcomer Granules
Some granules are highly mobile and sensitive to Ca²⁺ signals.
Two Types of Granules Contribute to Secretion:
1st Phase (Fast Response) → Docked granules fuse with the membrane immediately when Ca²⁺ increases.
2nd Phase (Sustained Release) → Some “restless newcomer granules” move directly to the membrane and get released, even though they weren’t docked earlier!
==> Some granules are highly mobile and can quickly move toward the membrane when needed.
This is important because β-cells can’t rely only on pre-docked granules—there aren’t enough of them for long-term insulin secretion.
The movement of these granules is controlled by microtubules & F-actin, which help them reach the membrane fast.
How are Insulin SGs Prioritised for Secretion?
Granule Age (New vs. Old Insulin)
Younger granules (newly synthesized insulin) are preferentially secreted over older, stored granules.
Experiment Evidence:
The study shows that islets release more newly labeled insulin (after 1-hour labeling) compared to older insulin (after 3-day labeling).
Fresh Insulin is More Effective
- Newer insulin is often better processed and packaged for secretion.
- Older granules might have structural or functional changes that make them less responsive to signals.
Some do get released during prolonged secretion.
Others might be recycled or broken down to maintain efficiency.
If β-cells become dysfunctional (like in diabetes), they might accumulate too many old granules, leading to insulin secretion defects.
How the Cytoskeleton Controls Insulin Granule Movement & Secretion in β-Cells
The Cytoskeleton Directs Granule Transport
1️⃣ Microtubules = The Highways (Green in Images)
Granules start deep inside the cell and need to travel toward the plasma membrane.
They ride along microtubules like cargo trucks heading to their destination.
Function: Act like highways that transport insulin granules from deep inside the β-cell toward the plasma membrane (PM).
Structure: Made of tubulin proteins, forming long, rigid tracks for movement.
Motor Proteins: Granules attach to motor proteins (kinesin, dynein) that “walk” along the microtubules, pulling granules toward the PM for secretion.
2️⃣ F-Actin = The Barrier at the Exit Ramp (Red in Images)
Function: Regulates access to the plasma membrane.
This acts like a toll gate or fence, blocking granules from getting out until the right signal (glucose increase) arrives.
At rest, F-actin forms a barrier, blocking insulin granules from reaching the PM.
When glucose increases, F-actin reorganizes, allowing granules to pass through and be released.
Structure: Made of actin filaments, forming a dense meshwork at the cell cortex (near the membrane).
Think of it like:
Microtubules = Highways (move granules forward).
F-actin = Toll gates (control when granules actually reach the destination).
How does this work?
At rest (low glucose), granules are mostly stored in deeper pools inside the β-cell.
When glucose increases, β-cells mobilize granules to the plasma membrane (PM).
for insulin release.
This requires microtubules to transport granules and F-actin to reorganize, removing its barrier function.
What Happens to Older Granules?
Over time, granules that aren’t released accumulate in F-actin-rich regions.
These granules are either:
✅ Slowly secreted during prolonged stimulation
❌ Recycled or disposed of in actin-positive multigranular bodies (like a cellular “trash system” for old granules).
1️⃣ How Do We Track Insulin Granule Age?
A fluorescent timer protein (dsRedE5TIMER) was used to label insulin granules over time.
When granules first form, they appear green.
As they age, they shift to yellow, then red.
This allows researchers to visually track how long granules have been in the cell.
Key Insight: By tracking color changes, scientists can determine whether younger or older granules are being released.
2️⃣ Identifying Aging Granules by Fluorescence Sorting
Flow cytometry was used to separate young (green) and old (red) granules at different time points (18h, 24h, 48h, 72h).
The number of old granules increases over time, showing that some insulin granules persist in the cell instead of being released.
Key Finding: Older granules accumulate over time, while younger granules are more likely to be secreted.
1️⃣ Older Granules Associate More with F-Actin – Why Does This Matter?
🔬 Observation:
Researchers found that older insulin granules tend to be more closely associated with F-actin filaments compared to younger granules.
This means that as granules age, they become more likely to be trapped in the F-actin network rather than being released.
🧬 Experiment & Evidence:
Using fluorescence tracking (dsRedE5TIMER), they labeled granules at different ages and monitored their movement.
Older granules were stuck near F-actin structures instead of freely moving toward the membrane.
🚨 Why Is This Important?
It explains why younger granules are preferentially secreted—they are less restricted by the F-actin barrier.
Older granules may be stored, recycled, or degraded instead of being released.
2️⃣ When Microtubules Are Disrupted (Nocodazole Treatment), Granule Movement Is Impaired
🔬 Observation:
Microtubules act as highways for granule transport.
When microtubules were disrupted using nocodazole (a drug that breaks down microtubules), granules could not move properly toward the membrane.
🧬 Experiment & Evidence:
In normal cells, granules moved smoothly along microtubules toward the membrane.
After treating cells with nocodazole, granules became disorganized and stuck inside the cell.
Fewer granules reached the plasma membrane, leading to reduced insulin secretion.
🚨 Why Is This Important?
This confirms that microtubules are essential for delivering granules to the membrane.
Without microtubules, insulin secretion fails because granules can’t be transported properly.
When F-Actin Is Disrupted (Latrunculin A Treatment), More Granules Reach the PM
🔬 Observation:
F-actin normally acts as a barrier, blocking granules from reaching the plasma membrane (PM).
When F-actin was disrupted using latrunculin A, granules moved more freely and more insulin was secreted.
🧬 Experiment & Evidence:
In normal conditions, only some granules could pass through the F-actin barrier to be secreted.
After treating β-cells with latrunculin A, the F-actin barrier was removed, and a larger number of granules reached the PM.
This led to a stronger insulin release response.
🚨 Why Is This Important?
It confirms that F-actin functions as a gatekeeper—it controls how many granules can be secreted at a time.
If F-actin doesn’t reorganize properly in response to glucose, granules get stuck and insulin secretion is impaired (which could contribute to diabetes).
wording of microtubule transport of SG
1️⃣ SGs Travel on Microtubules Toward the Actin-Rich Cortex
“During maturation, SGs undergo microtubule-dependent trafficking away from the inner cellular radius and are passed to the actin-rich cortex.”
🔬 What this means:
Newly made insulin granules start deep inside the β-cell near the Golgi.
They travel outward toward the plasma membrane (PM) using microtubule “highways.”
The outer region of the cell (near the PM) is rich in F-actin, forming a dense network known as the actin cortex.
When SGs reach this region, they pause at the F-actin barrier, waiting for the signal to be released.
🧬 Key point:
✅ Microtubules bring granules to the edge of the cell, but F-actin controls whether they can be released.
wording of F-actin block
2️⃣ In Resting (Basal) Conditions, F-Actin Blocks Insulin Release
“In basal conditions, F-actin functions as a barrier to block SNARE-complex formation.”
🔬 What this means:
At rest (low glucose levels), F-actin forms a dense barrier that prevents SGs from reaching the membrane.
The SNARE complex is a set of proteins (including VAMP2, Syntaxin, and SNAP-25) needed for exocytosis (insulin release).
If SNAREs can’t form, SGs can’t fuse with the PM, and insulin isn’t released.
🧬 Key point:
✅ F-actin is a gatekeeper—it prevents unnecessary insulin secretion when glucose is low.
