Culture Collections

New cell lines for autophagy research

ULK cell lines for autophagy research - HOMEPAGE

 

A series of mouse embryonic fibroblasts with different capability for uncoordinated (Unc)-51-like kinase 1 and 2 (ULK1 and ULK2) activity, which are known to play a critical role during the activation of autophagy, are now available from Public Health England’s European Collection of Authenticated Cell Cultures (ECACC).  Autophagy is the mechanism by which a cell degrades unnecessary or dysfunctional cellular components and has been implicated in several medical scenarios such as cancer, neurodegeneration and immunity related disorders.  The cell lines were deposited by Dr Sharon Tooze of Cancer Research UK.

 

Mouse embryonic fibroblasts for studying autophagy with different ULK1 and ULK2 statuses:

 

Catalogue No.

Cell line name

ULK 1 and 2 status         

Immortalisation method             

14050808

ULK1/2 WT MEF (SIM)

 

Wild type

 

Spontaneously immortalised

 

14050804

 

ULK1/2 WT MEF (SV40)

 

Wild type

 

SV40-immortalised

 

14050806

 

ULK1 KO MEF (SIM)

 

ULK1 knock-out

 

Spontaneously immortalised

 

14050807

 

ULK1 KO MEF (SV40)

 

ULK1 knock-out

 

SV40-immortalised

 

14050810

 

ULK2 KO MEF (SIM)

 

ULK2 knock-out

 

Spontaneously immortalised

 

14050805

 

ULK2 KO MEF (SV40)

 

ULK2 knock-out

 

SV40-immortalised

 

14050803

 

MEF Ulk1 -/- Ulk2 -/- (DKO) (SIM)

 

Double knock-out

 

Spontaneously immortalised

 

14050802

 

MEF Ulk1 -/- Ulk2 -/- (DKO) (SV40)

 

Double knock-out

 

SV40-immortalised

 

 

Autophagy targets long-lived cellular proteins, organelles, ubiquitinated cellular substrates and aberrant proteins. During autophagy in mammalian cells membrane cisterna engulf cytoplasmic materials to form nascent autophagosomes, which then fuse with endosomal structures to initiate the degradation of their contents.

The serine-threonine protein kinase Atg1 has been identified as a key regulator of autophagy in yeast (1, 2, 3).  ULK1 and ULK2 are serine/threonine protein kinases, the mammalian homologues of the yeast Atg1 kinase, which is known to play a critical role during the activation of autophagy. Studies have demonstrated that the conserved C-terminal domain (CTD) of ULK1 controls the regulatory function and localisation of the protein.

The research group led by Dr Sharon Tooze of Cancer Research UK has explored the role of ULK kinase activity and intra-molecular interactions to further understand their function and demonstrated that the dominant-negative activity of kinase-dead mutants requires a 7-residue motif within the CTD. They have developed a model in which the functions of ULK1 and ULK2 are controlled by autophosphorylation and conformational changes involving exposure of the CTD (4, 5).

This same group have established a series of mouse embryonic fibroblast cell lines isolated from mice that are wild-type, homozygous or double knock-outs at the ULK1 & ULK2 loci. These cell lines have been established by spontaneous immortalisation (SIM) or by transformation with SV40.

 

References

1. Matsuura A, Tsukada M,  Wada Y, and Ohsumi Y.  Apg1p, a novel protein kinase required for the autophagic process in Saccharomyces cerevisiae. Gene 1997,192:245–250. PMID: 9224897.

2. Straub M, Bredschneider M, and Thumm M. AUT3, a serine/ threonine kinase gene, is essential for autophagocytosis in Saccharomyces cerevisiae. J. Bacteriol. 1997,179:3875–3883. PMID: 9190802  

3. Thumm, M., Egner R, Koch B, Schlumpberger M, Straub M, Veenhuis M, and Wolf DH. Isolation of autophagocytosis mutants of Saccharomyces cerevisiae. FEBS Lett. 1994 349:275–280, PMID: 8050581

4. Chan EY, Longatti A, McKnight NC, Tooze SA. Kinase-inactivated ULK proteins inhibit autophagy via their conserved C-terminal domains using an Atg13-independent mechanism. Mol Cell Biol. 2009 Jan;29 (1):157-71. doi: 10.1128/MCB.01082-08. Epub 2008 Oct 20. PMID: 18936157

5. McAlpine F, Williamson LE, Tooze SA, Chan EY. Regulation of nutrient-sensitive autophagy by uncoordinated 51-like kinases 1 and 2. Autophagy. 2013 Mar;9(3):361-73. doi: 10.4161/auto.23066. Epub 2013 Jan 4. PMID: 23291478

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