92
distinct subfamilies B/PPP2R2, B ́/PPP2R5, B ́ ́/PPP2R3 and B ́ ́ ́/Striatins.
Alternative splicing of some of the mRNAs gives rise to at least 23 different B-type
subunits. Theoretically, the combinatorial assembly of one B-type subunit with one
A- and C-subunit would yield more than 90 different PP2A holoenzymes (Sents
et al. 2013 ). Mouse studies revealed that knockout of the PPP2CA gene is embry-
onic lethal despite the presence of the β-isoform (PPP2CB) which shares 97 %
sequence identity with the α-isoform (Gotz et al. 1998 ). Thus, even the two C-subunit
isoforms seem to have nonredundant functions despite their high levels of sequence
identity, indicating that the 90 different possible holoenzymes have different sub-
strate specificities, regulations and/or localisations. Notably, PP2As can undergo a
complex and highly regulated maturation process. To date, the best understood post-
translational modifications of PP2A include C-terminal methylation and phosphor-
ylation of the catalytic subunit. Both modifications are involved in a surveillance
mechanism that controls PP2A holoenzyme assembly and prevents premature
unspecific activity of the catalytic subunit (Stanevich et al. 2011 ; Hombauer et al.
2007 ). An additional mechanism controlling the activity of PP2As involves the
binding of inhibitory proteins. CIP2A, SET/I2PP2A and Anp32/I1PP2A are inhibitor
proteins that decrease the activity of PP2A against a broad range of substrates (Li
et al. 1995 ; Junttila et al. 2007 ; Laine et al. 2013 ). Their importance is highlighted
by the fact that upregulation of these inhibitors and the resulting inhibition of PP2A
activity are associated with the development of tumours, e.g. by increasing the sta-
bility of the oncogene Myc (Junttila et al. 2007 ; Neviani et al. 2005 ).
3.5.1 The PP2A-B55 Phosphatase
The complete identity of the phosphatases involved in maintaining the dephosphor-
ylated interphase state and/or reverting the mitotic phosphorylated state to the
dephosphorylated state during exit from M-phase and their relative contribution in
these processes remains elusive. However, experiments in invertebrates and verte-
brates clearly identify PP2A with a regulatory B55 subunit (PP2A-B55) as a key
antagonist of Cdk1/cyclin-B (Fig. 3.5) (Mochida et al. 2009 ; Manchado et al. 2010 ;
Schmitz et al. 2010 ; Cundell et al. 2013 ; Mayer-Jaekel et al. 1994 ). siRNA- mediated
knockdown of B55 in cells or immunodepletion of B55 from cytoplasmic extracts
blocks dephosphorylation of several Cdk1 model substrates and, with varying effi-
ciency, general dephosphorylation of mitotic phosphoproteins.
3.5.1.1 Regulation of PP2A-B55
Xenopus laevis egg extract studies revealed that the activity of PP2A-B55 is regu-
lated reciprocally to that of Cdk1, i.e. it is downregulated at mitotic entry and reac-
tivated at the beginning of mitotic exit (Mochida et al. 2009 ). This modulation of
PP2A-B55 activity depends on the activity of Cdk1. How is Cdk1 able to inactivate
A. Heim et al.