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STUDY OF TRPML CHANNELS REVEALS INSIGHT INTO ENDOCYTIC MALFUNCTION, ORGANELLE CROSSTALK, AND THE ACTIVATION OF PRO-APOPTOTIC PATHWAYS

Colletti, Grace (2012) STUDY OF TRPML CHANNELS REVEALS INSIGHT INTO ENDOCYTIC MALFUNCTION, ORGANELLE CROSSTALK, AND THE ACTIVATION OF PRO-APOPTOTIC PATHWAYS. Doctoral Dissertation, University of Pittsburgh. (Unpublished)

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Abstract

Mucolipidosis type IV (MLIV) is a lysosomal storage disease resulting from mutations in the gene MCOLN1, which codes for a transient receptor potential family ion channel TRPML1 (Mucolipin-1). MLIV has an early onset and is characterized by developmental delays, motor and cognitive deficiencies, gastric abnormalities, retinal degeneration and corneal cloudiness. The degenerative aspects of MLIV have been attributed to cell death, whose mechanisms remain to be delineated in MLIV and most other lysosomal storage diseases. The function of TRPML1 is still not completely understood in the cell. In order to address the function of this channel as well as the consequences of its loss, we have studied TRPML1 and a closely related channel TRPML3 using proteomic, transcriptional, and ion channel activity assays. Structure/function analyses of TRPML (Mucolipin) channels revealed that the closely related TRPML3 channel conducts K+, Na2+, Ca2+, and Fe2+, but does not conduct transition metals such as Cu2+ and Co2+. The permeability of the channel can be disrupted by mutations to the pore domain and a constitutively active channel mutation abolishes transition metal block of both TRPML1 and TRPML3. The similarities in TRPML sequence and the activity of the constitutively active mutants suggests that these channels may share similar activity profiles, which supports previous reports that TRPML1 may function to regulate lysosomal Ca2+ and Fe2+ in order to promote proper lysosomal fission/fusion events and function. Our proteomic and transcriptional analyses reveal that acute downregulation of TRPML1 results in apoptosis in a cathepsin B and Bax dependent manner. This is the first evidence that acute TRPML1 loss is linked to apoptosis and my work provides a preliminary mechanism for this process. Furthermore, TRPML1 loss results in increased NFκB levels as well as altered transcription of genes outside the previously identified lysosomal gene expression network. This suggests that loss of TRPML1 results in cellular changes that affect downstream transcriptional targets and induces cellular responses, such as apoptosis. Taken together, these data shed light on the importance of TRPML1 function and may help us understand the cellular role of this channel and how its loss results in cell death.


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Details

Item Type: University of Pittsburgh ETD
Status: Unpublished
Creators/Authors:
CreatorsEmailPitt UsernameORCID
Colletti, Gracegac13@pitt.eduGAC13
ETD Committee:
TitleMemberEmail AddressPitt UsernameORCID
Committee ChairKirill, Kiselyovkiselyov@pitt.eduKISELYOV
Committee MemberHildebrand, Jeffreyjeffh@pitt.eduJEFFH
Committee MemberOra, Weiszweisz@pitt.eduWEISZ
Committee MemberMichael , Grabemdgrabe@pitt.eduMDGRABE
Committee MemberWilliam, Saunderswsaund@pitt.eduWSAUND
Date: 31 January 2012
Date Type: Publication
Defense Date: 29 November 2011
Approval Date: 31 January 2012
Submission Date: 8 December 2011
Access Restriction: No restriction; Release the ETD for access worldwide immediately.
Number of Pages: 147
Institution: University of Pittsburgh
Schools and Programs: Dietrich School of Arts and Sciences > Biological Sciences
Degree: PhD - Doctor of Philosophy
Thesis Type: Doctoral Dissertation
Refereed: Yes
Uncontrolled Keywords: Mucolipin-1, TRPML1, Mucolipidosis Type IV, Cathepsin B mediated apoptosis, Endocytic malfunction, Transition metal toxicity, TRPML3
Date Deposited: 31 Jan 2012 16:47
Last Modified: 15 Nov 2016 13:55
URI: http://d-scholarship.pitt.edu/id/eprint/10738

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  • STUDY OF TRPML CHANNELS REVEALS INSIGHT INTO ENDOCYTIC MALFUNCTION, ORGANELLE CROSSTALK, AND THE ACTIVATION OF PRO-APOPTOTIC PATHWAYS. (deposited 31 Jan 2012 16:47) [Currently Displayed]

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