of the Max - MDC

Structure of the Group
Group Leader
Prof. Dr. Ingo Morano
Scientists
Dr. Hannelore Haase
Dr. Daria Petzhold
Dr. Andreas Marg
Graduate and
Undergraduate Students
Lars Schulz
Christiane Look
Romy Siegert
Janine Lossie
Nicole Bidmon
Ines Pankonien
Ivonne Heisse
Dana Rotte
Technical Assistants
Petra Pierschalek
Steffen Lutter
Wolfgang-Peter Schlegel
Mathias Pippow
Secretariat
Manuela Kaada
wild-type and knock-out animals were similar. Thus, initial
phasic contraction is generated by SM-MyHC recruitment
while the sustained tonic contraction state can be produced
by NM-MyHC activation, which represent the latch crossbridges
in smooth muscle (Morano et al. 2000, Nature Cell
Biology, 2, 371–375, Löfgren et al. 2003 J. Gen. Physiol.
12, 301-310).
Ahnak is a player in the sympathetic control of
the cardiac L-type calcium channel
Sympathetic tone is a major determinant of the L-type Ca 2+
channel activity, thus regulating influx of Ca 2+ from exterior
into the cytosol of cardiomyocytes (I CaL ). Sympathetic stimulation
of I CaL is known to be mediated by a cascade of reactions
involving beta-adrenergic receptors, G-protein coupled
adenylyl cyclase, and protein kinase A (PKA). The
molecular basis of this regulation, in particular the site(s)
targeted by PKA, as well as the mechanism by which phosphorylation
increased I CaL remained obscure. In fact, the
postulated link between Cav1.2 phosphorylation and
enhanced I CaL could not be demonstrated in the Xenopus
oocyte expression system. Thus, PKA-dependent regulation
of I CaL may require additional unidentified components.
Searching for those “missing links“ in mammalian cardiomyocytes
led us to the identification of the 700-kDa ahnak protein,
which was initially characterized by coprecipitation
with the Cavβ2 subunit and in vivo phosphorylation in
response to sympathetic stimulation of the heart (Haase et
al. 1999; Faseb J. 13:2161-72).
In the heart, ahnak is expressed in cardiomyocytes,
endothelial cells, and smooth muscle cells. At the subcellular
level, ahnak locates to the cytoplasmic aspect of the sarcolemma
in cardiomyocytes (Figure 2). It interacts with the
channel β2-subunit via multipoint attachment mediated by
ahnak´s carboxy-terminal domains, ahnak-C1 and ahnak-
C2. The most C-terminal ahnak-C2 domain has actin-binding
and actin-bundling capacity. As such it provides a link to
the subsarcolemma cytoskeleton and stabilizes muscle contractility
(Haase et al. 2004, Faseb J. 18:839-42). Patchclamp
experiments on rat ventricular cardiomyocytes
showed that targeting the high affinity ahnak-C2/ β2-subunit
interaction by a peptide competition approach leads to
an increase in the Ca 2+ current amplitude and a slowing of
channel inactivation (Alvarez et al. 2004, J Biol Chem.
279:12456-61). These results suggested that endogenous
ahnak exerts a sustained inhibitory effect on I CaL by strong
β2-subunit binding via the ahnak-C2 domain. Furthermore,
the interaction between ahnak-C1 and β2-subunit plays a
critical role for the sympathetic regulation of L-type Ca 2+
channel activity: PKA phosphorylation reduced the interaction
between ahnak-C1 and the β2-subunit, thus releasing
its inhibitory effect on I CaL (Haase et al. 2005).
We screened a patient cohort with hypertrophic cardiomyopathy
in order to identify naturally occurring, genetic
ahnak variants. The identification of the coding genetic
variant Ile5236Thr-ahnak prompted us to study functional
consequences of this mutation on β2-subunit binding and
Ca 2+ channel function. We found that Ile5236Thr ahnak
interfered with the classic beta-adrenergic regulation of I CaL
(Figure 3)
Selected Publications
Woischwill, Ch, Karczewski, P, Bartsch, H, Luther, HP, Kott, M,
Haase, H, Morano, I. (2005). Regulation of the human atrial
myosin light chain 1 promoter by Ca 2+ -calmodulin-dependent
signalling pathways. FASEB J. 19, 503-511
Haase, H, Alvarez, J, Petzhold, D, Doller, A, Behlke, J, Erdmann,
J, Hetzer, R, Regitz-Zagrosek, V, Vassort, G, Morano, I. (2005).
Ahnak is critical for cardiac calcium calcium channel function
and its beta-adrenergic regulation. FASEB J. 19, 1969-1977
Haase, H, Dobbernack, G, Tünnemann, G, Karczewski, P,
Cardoso, C, Petzhold, D, Schlegel, WP, Lutter, St, Pierschalek, P,
Behlke, J, Morano, I. (2006). Minigenes encoding N-terminal
domains of human cardiac myosin light chain-1 improve heart
function of transgenic rats. FASEB J. 20, 865-873
Tünnemann, G, Behlke, J, Karczewski, P, Haase, H. Cardoso,
MCh, Morano, I. (2007). Modulation of muscle contraction by
a cell permeable peptide. J. Mol. Med. In press
Aydt Wolff Morano (2007) Molecular modeling of the
myosin-S1(A1) isoform. J Struct Biol., 159,158-63.
Haase H (2007) Ahnak, a new player in β-adrenergic regulation
of the cardiac L-type Ca2+ channel. Cardiovasc Res., 73,19-25.
40 Cardiovascular and Metabolic Disease Research