Human telomerase exists in two distinct active complexes in vivo

Area of Science:

• Molecular Biology
• Biochemistry
• Cell Biology

Background:

• Telomerase maintains telomere length, crucial for cellular aging and cancer.
• Human telomerase comprises telomerase reverse transcriptase (TERT) and template RNA (TERC).
• Understanding telomerase complex composition and regulation is vital for therapeutic development.

Purpose of the Study:

• To identify and characterize distinct active human telomerase complexes.
• To investigate the role of Hsp90 in telomerase complex formation and activity.
• To elucidate the stability and regulation mechanisms of human telomerase complexes.

Main Methods:

• Purification of recombinant human telomerase reconstituted in insect cells.
• Biochemical assays to assess telomerase activity in vitro.
• Analysis of telomerase complexes in HeLa cells using affinity purification and Western blotting.
• Investigation of proteasomal degradation pathways.

Main Results:

• Two active human telomerase complexes (680 kDa and 380 kDa) were identified in insect cells.
• The 680 kDa complex lacks Hsp90 and is resistant to Hsp90 inhibitors.
• The 380 kDa complex contains Hsp90, is sensitive to inhibitors, and undergoes proteasome-mediated degradation of hTERT.
• Similar complexes with corresponding properties were found in HeLa cells.

Conclusions:

• Human telomerase exists in at least two distinct active complexes with differing properties.
• Hsp90 association influences telomerase complex stability and sensitivity to inhibitors.
• Proteasome-mediated degradation regulates the abundance of the Hsp90-containing telomerase complex.