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Localization of serum-derived alpha 2 macroglobulin in cultured cells and decrease after Moloney sarcoma virus

I Pastan, M Willingham, W Anderson

|November 1, 1977

Summary

This summary is machine-generated.

Cultured fibroblasts absorb alpha 2 macroglobulin (alpha 2M) from serum, storing it in vesicles distinct from lysosomes. This protease inhibitor may influence cell behavior by inhibiting cellular proteases.

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Area of Science:

Cell biology
Biochemistry
Protease inhibition

Background:

Cultured fibroblasts, including NRK cells, contain alpha 2 macroglobulin (alpha 2M).
Alpha 2M is acquired from calf serum present in the cell culture medium.
A portion of the internalized alpha 2M is subsequently released back into the medium.

Purpose of the Study:

To investigate the intracellular localization and origin of alpha 2 macroglobulin (alpha 2M) in cultured fibroblasts.
To determine if alpha 2M synthesized by cells is released into the culture medium.
To explore the potential role of alpha 2M in mediating serum effects on cell behavior.

Main Methods:

Radiolabeling experiments using 14C-amino acids to trace protein synthesis.
Fluorescence microscopy with rhodamine-labeled antibodies to visualize alpha 2M in fixed cells.
Fluorescence microscopy of living cells to observe the uptake of rhodamine-labeled alpha 2M.

Main Results:

Intracellular and extracellular alpha 2M showed no detectable radioactivity, indicating it is not synthesized by the cells.
Alpha 2M was localized in vesicular organelles distinct from primary lysosomes.
Rhodamine-labeled alpha 2M was observed to be taken up into similar vesicular structures in living cells.
Moloney sarcoma virus-transformed cells exhibited the lowest levels of alpha 2M.

Conclusions:

The alpha 2 macroglobulin found in cultured fibroblasts originates from the culture medium's serum.
Alpha 2M is sequestered within specific intracellular vesicles, not lysosomes.
Serum-derived alpha 2M may modulate cellular functions through protease inhibition, potentially affecting cell behavior.