TFEB Localization and MK-STYX Expression in HeLa Trials 1 & 2

In order to determine the changes in TFEB (transcription factor EB) localization within HeLa cells in the presence of the pseudophosphatase MK-STYX, HeLa cells were transfected with pMT2 (control), MK-STYX, and F1 (active mutant of MK-STYX). After 24 hours, transfected cells were serum-starved over night (~18 hours) and immunostained for the TFEB, an autophagy regulator. TFEB is normally localized throughout the cell, including the cytoplasm and the nucleus, but its presence in the nucleus would indicate an upregulation of autophagy. After running the first two trials, fluorescent microscopy results indicated a role of MK-STYX in inducing TFEB localization. However, the data for the cells transfected with pMT2 could not be used because most of these cells lysed after immunostaining. At this point, I suspect that the cells might have lysed due to dehydration during immunostaining, being overstressed, or both.

Comments

  1. ammattei says:

    Thank you for the update! I can’t wait to see how this analysis of TFEB, the master regulator of autophagy, turns out. If your preliminary findings are verified by your upcoming trials, an exciting array of experiments investigating the proteins and organelles under the control of TFEB will be opened up. Keep up the good work!

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