MINIREVIEW
Huntington’s disease: roles of huntingtin-interacting
protein 1 (HIP-1) and its molecular partner HIPPI in the
regulation of apoptosis and transcription
Nitai P. Bhattacharyya, Manisha Banerjee and Pritha Majumder*
Crystallography and Molecular Biology Division and Structural Genomics Section, Saha Institute of Nuclear Physics, Kolkata, India
Huntington’s disease (HD, OMIM 143100) is an auto-
somal dominant progressive neurodegenerative disease
caused by the expansion of polymorphic CAG (coding
for glutamine) repeats beyond 36 at exon 1 of the
huntingtin (htt) gene, localized at chromosome 4p16.3.
Age at onset (AO) of the disease varies widely
(1–90 years, mean  35 years). There is an inverse cor-
relation between AO and expanded CAG repeat
numbers, but it is not the only determinant of
variation in AO [1]. HD is fatal within 10–15 years
after appearance of the first symptom. The symptoms
include uncontrolled movement, emotional distur-
bances, psychiatric abnormalities, cognitive deficits,
and dementia. The gene htt encodes a protein [hunting-
tin (Htt),  348 kDa] with a polyglutamine stretch
starting from the 18th amino acid. Also, two proline-
rich regions adjacent to the polyglutamine domain and
several HEAT repeats, known to be involved in
Keywords
apoptosis; HIP-1; HIPPI; huntingtin-
interacting proteins; transcription
Correspondence
N. P. Bhattacharyya, Crystallography and
Molecular Biology Division and Structural
Genomics Section, Saha Institute of Nuclear

D-aspartate receptor; pDED, pseudo-death effector
domain; Shh, Sonic hedgehog.
FEBS Journal 275 (2008) 4271–4279 ª 2008 The Authors Journal compilation ª 2008 FEBS 4271
protein–protein interactions, are present at the N-ter-
minal region of the protein. Wild-type Htt is localized
at the endoplasmic reticulum, Golgi complex, mito-
chondria, and synaptic vesicles. Htt is ubiquitously
expressed, although the neurodegeneration caused by
the mutated Htt shows region specificity [2,3].
The expanded polyglutamine domain of mutant Htt
is highly self-associative, resulting in aggregates ⁄ neuro-
nal intranuclear inclusions. Aggregates ⁄ neuronal intra-
nuclear inclusions are observed in cell models, brains
of transgenic animals, and post-mortem brains of HD
patients [2]. Aggregate formation is enhanced with the
increase in the number of glutamines in vitro and
in vivo, and is believed to cause neurodegeneration [4].
Although a contradictory finding, that visible aggre-
gates are protective to neurons, has also been made
[5]. The autosomal dominant nature of the disease sug-
gests a toxic gain-of-function of the mutated protein
that disrupts normal cellular functions and causes
neuronal death [3]. Loss-of-function of the wild-type
protein may also contribute, at least partially, to the
disease pathology [6]. Over the years, various cellular
events, such as excitotoxicity, oxidative stress, mito-
chondrial dysfunction, stress in the endoplasmic reticu-
lum, formation of channels through membranes,
axonal transport, protein degradation, autophagy,
transcriptional dysregulation, and apoptosis, have been

chain A (CLTA), and clathrin light chain B (CLTB),
and N-methyl-d-aspartate receptor (NMDAR) subun-
its NR2A and NR2B. Various domains, such as the
AP180 N-terminal homology domain (ANTH), also
known as the Epsin N-terminal homology (ENTH)
domain, the central coiled-coil region and a C-terminal
talin homology domain are present at HIP-1. The
coiled-coil domain contains a leucine-zipper motif and
mediates heterodimerization with HIP-1R. Consensus
binding sites for the endocytic adaptor protein AP2
(DPF motif), clathrin heavy chain (LMDMD clathrin-
box motif) and a phosphatidylinositol 4,5-biphosphate-
binding motif at its ANTH ⁄ ENTH domain are also
present [14,18]. Various domains of HIP-1 are shown
in Fig. 1. Direct evidence that HIP-1 is involved in
endocytosis comes from HIP-1 knock-out (HIP-1
) ⁄ )
)
mice, which show defects in assembly of endocytic
protein complexes on liposomal membranes and
a-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid
receptor trafficking [19]. The similarities in amino acid
sequences and domains between HIP-1, HIP-1R and
yeast ortholog Sla2p, the interacting partners of HIP-1
with known functions and results with knockout mice
Fig. 1. Various domains of HIP-1. The ANTH ⁄ ENTH domain (38–160), coiled-coil domain (371–610), and talin-like domain (814–1112) were
predicted with the
SMART tool ( Binding sites for HIPPI (422–503), AP2 (262–266 and 358–360), CLH1, CLH2
(332–336), CLTA, CLTB (484–489) and other domains are taken from the published literature and mentioned in the text. The positions of the
amino acids are not to scale.

tosis [17,21]. Wild-type N-terminal Htt, being able to
interact with HIP-1 strongly, may reduce the amount
of HIP-1 that is available to interact with other pro-
tein(s) and reduce apoptosis. In rat cells, HIP-1 is
cleaved in response to drugs that are known to induce
apoptosis, as well as in cells expressing exogenous
HIP-1, although the relevance of such cleavages in
apoptosis remains unknown. HIP-1 interacts directly
with procaspase-9 and activates it. Direct interaction
of HIP-1 with Apaf1 increases recruitment of cyto-
chrome c to the apotosome complex, resulting in
increased apoptosis [21]. Depending on the status of
phosphorylation of HIP-1 by Dyrk1, HIP-1 interacts
with caspase-3 and enhances apoptosis, in a condition
where interaction and phosphorylation of HIP-1 by
Dyrk1 are reduced [22].
Exogenous expression of HIPPI (HIP-1 protein
interactor), a molecular partner of HIP-1, increases
apoptosis through the extrinsic pathway. The HIP-1–
HIPPI heterodimer recruits procaspase-8 and activates
it [23]. Enhancement of apoptosis by exogenous HIPPI
in the presence of endogenous HIP-1 is mediated
through activation of caspase-8, caspase-1, caspase-9 ⁄
caspase-6, and caspase-3. Cleavage of Bid and release
of cytochrome c and apoptosis-inducing factors from
the mitochondria are also observed. Coexpression of
wild-type htt exon 1 and Hippi decreases apoptosis and
increases survival in comparison with that obtained in
cells expressing Hippi only. In such a condition, inter-
action of HIPPI with HIP-1 is reduced. This result fur-

mentioned earlier [25]. Taken together, these results
show that HIP-1 may act as a prosurvival protein in
different conditions.
Contradictory results showing that HIP-1 is a proa-
poptotic protein [17,21–24] or an antiapoptotic protein
[25–30] in different conditions could be due to the
presence or absence of HIP-1-interacting partners. The
decrease in HIP-1–HIPPI-mediated apoptosis, either
by overexpression of Homer 1c, an interactor of HIP-
PI (for details see the next section), or by the wild-type
N-terminal Htt, which strongly interacts with HIP-1
[24,31], supports this contention. In such cases, the
amounts of freely available HIPPI or HIP-1 may
decrease, resulting in reduced apoptosis. Exogenous
expression of HIP-1 (cloned in pcDNA3 and kindly
provided to us by T. S. Ross, Department of Internal
Medicine, University of Michigan Medical School,
Ann Arbor, MI, USA) in HeLa cells, where endoge-
N. P. Bhattacharyya et al. HIP-1 & HIPPI mediated apoptosis and transcription
FEBS Journal 275 (2008) 4271–4279 ª 2008 The Authors Journal compilation ª 2008 FEBS 4273
nous HIPPI is undetectable [24], did not increase
apoptosis. However, in Neuro2A and K562 cells,
where endogenous HIPPI is present [24], exogenous
expression of HIP-1 increased apoptosis. Additionally,
expression of HIPPI in HeLa cells, where HIP-1 was
knocked down, decreased apoptosis (M. Banerjee and
N. P. Bhattacharyya, unpublished observations) rela-
tive to that obtained in HeLa cells with endogenous
HIP-1 [24]. The proapoptotic activity of HIPPI or
HIP-1 may thus be dependent on the presence or

with HIPPI and increase HIPPI-mediated apoptosis
through the caspase-8-mediated pathway. Rybp also
interacts with ubiquitin-binding protein, procaspase-8,
procaspase-10, and the HIPPI interactor apotin. Inter-
action of HIPPI with Rybp is involved in murine
neural development, although the significance of such
an interaction in apoptosis regulation or HD patho-
genesis remains elusive [33]. Apotin, a chicken anemia
virus-encoded protein, has been shown to colocalize
with HIPPI in the cytoplasm of normal cells, whereas
in tumor cells, they localize separately in the nucleus
and cytoplasm. The HIPPI–apoptin interaction may
suppress apoptosis [35]. The functional relevance of
such interactions in HIP-1- or HIPPI-mediated apop-
tosis also remains unknown. Biogenesis of lysosome-
related organelles complex-1 subunit 2 (BLOC1S2)
specifically interacts with HIPPI, but not with HIP-1.
Coexpression of HIPPI and BLOC1S2 does not increase
apoptosis but sensitizes apoptosis induction by stauro-
sporin or death ligand. In addition, the expression of
BLOC1S2 is increased in some tumors [34].
Information on the interacting partners of HIPPI
such as HIP-1, Homer 1c, BAR and apoptin
indicates that HIPPI may also be a proapoptotic
protein. Rybp, an interactor of HIPPI, interacts with
caspase-8 and caspase-10, indicating that HIPPI
might also be involved in the regulation of apoptosis.
Even though the exact function of HIPPI remains
unknown, knockout mice for Hippi (HIPPI
) ⁄ )

prosurvival ⁄ antiapoptotic functions described in the
preceding sections.
The evidence that HIPPI directly or indirectly alters
gene expression comes from the observations that
caspase-1, caspase-3, caspase-7, caspase-8 and caspase-10
expression is increased in cells expressing exogenous
HIP-1 & HIPPI mediated apoptosis and transcription N. P. Bhattacharyya et al.
4274 FEBS Journal 275 (2008) 4271–4279 ª 2008 The Authors Journal compilation ª 2008 FEBS
Hippi. Also, expression of the mitochondrial-coded
genes ND1 and ND4, the nuclear genome-coded
mitochondrial genes SDHA and SDHB and the antia-
poptotic genes BCL-2 and survivin is decreased in
Hippi-expressing cells [24]. Decreased expression of
ND1, ND4, SDHA, SDHB and BCL-2 may cause
mitochondrial dysfunctions and contribute towards the
increased apoptosis by HIPPI as mentioned earlier.
Decreased expression of the antiapoptotic gene survivin
may also enhance apoptosis.
HIPPI interacts with the putative promoter of
caspase-1 in vitro and in vivo [39]. On the basis of
in vitro interactions of various mutants of the sequence
5¢-AAAGACATG-3¢ ()101 to )93) present at the
caspase-1 putative promoter sequence, where HIPPI can
bind, it has been predicted that HIPPI will interact with
AAAGA[GC][ATC][TG] [40]. The presence of other
sequence motifs around the HIPPI binding site where
transcription factors p53, p73 and ETS1 can bind and
influence the expression of caspase-1 [41–43] indicate
that these or other unknown transcription factors may
cooperate with HIPPI for the regulation of caspase-1

requires further confirmation. Rybp, also known as
death effector domain-associated factor, belongs to a
family of small zinc finger-containing proteins that
participate in transcriptional regulation by binding
with other transcription factors such as YY1 and E2F
or transcription repressors [44]. Additionally, the
Rybp-related protein Yaf2 interacts with HIPPI. Inter-
action of HIPPI with Rybp is proposed to be involved
in murine neural development, although the signifi-
cance of such an interaction in HD pathogenesis
remains elusive [33]. It is speculative that Rybp, a
molecular interactor of HIPPI, cooperates with HIPPI
to augment transcription of caspase-1, and this war-
rants further studies. A summary of the findings that
HIPPI increases apoptosis and alters gene expression is
shown in Fig. 3.
wH16-Hi
Pro-caspase-1
Caspase-1
Beta actin
45 kDa
20 kDa
42 kDa
Hi
Fig. 2. Western blot analysis for the expression of caspase-1 in HeLa cells expressing green fluorescent protein (GFP)-tagged HIPPI (GFP–
Hippi, lane denoted by Hi) and HeLa cells coexpressing GFP–HIPPI and the red fluorescent protein-tagged wild-type exon 1 of the htt gene
with 16 CAG repeats (DsRed–wH16, lane denoted by wH16-Hi). The lower panel shows the result with antibody to b-actin (42 kDa) as load-
ing control. The sizes of procaspase-1 (45 kDa) and the activated caspase-1 (20 kDa) are shown by the arrows on the left. The bar diagram
(right panel) shows the average (n = 3) of integrated optical density (IOD) of the bands obtained with antibody to caspase-1 in western blot
analysis using GFP–HIPPI-expressing cells (unfilled) and cells coexpressing GFP–HIPPI and DsRed–wH16 (filled bar).

tions. The mechanism by which caspase-1 expression is
increased in HD is not well understood, although the
protein is implicated in the progression of HD [47,48].
The regulation of caspase-1 by HIPPI observed in cul-
tured cells provides an explanation for the increased
caspase-1 expression in HD. In HD, owing to weaker
interactions of HIP-1 with the mutated Htt, the free
HIP-1 pool might increase, and this in turn would
lead to the formation of more HIP1–HIPPI, initiating
apoptosis by caspase-8 activation and its downstream
pathway, and might also increase the transcription of
caspase-1. Further studies using animal models are
necessary to confirm this.
HIPPI
Freely
available
HIP1
2
HIPPI
1
3
Strong
interaction
HIP1
6
4
5
Caspase-8
Nucleus
Weak

Different symbols representing different proteins are shown in the box. Numbers representing different processes are also shown.
HIP-1 & HIPPI mediated apoptosis and transcription N. P. Bhattacharyya et al.
4276 FEBS Journal 275 (2008) 4271–4279 ª 2008 The Authors Journal compilation ª 2008 FEBS
Conclusions
HIP-1 and its interacting partner HIPPI together
induce apoptosis by the intrinsic and extrinsic
pathways. Homer 1c, an interactor of HIPPI, and the
wild-type N-terminal Htt, which interacts strongly
with HIP-1, reduce HIP-1–HIPPI-mediated apoptosis.
In the presence of Homer 1c, HIPPI may interact
preferentially with it, resulting in a decrease of the
amount of HIP-1–HIPPI heterodimer and apoptosis
induction. The effect of HIP-1 may depend not only
on the amount of the interacting proteins but also on
the affinities of interacting proteins. The uncreased
expression of caspase-1 observed in HD may be medi-
ated through HIPPI. The role of HIP-1 in transloca-
tion of HIPPI into the nucleus and that of other
transcriptional regulators cooperating with HIPPI are
yet to be determined. If the observations in cell
culture are replicated in HD models or post-mortem
brains, an explanation of the increased expression of
caspase-1 and other subset of genes altered in HD
may be available.
Acknowledgements
We acknowledge Professors A. Mukherjee, D. Mukho-
padhyay and M. S. Moumita Datta for critically read-
ing the manuscript and their valuable suggestions.
References
1 Squitieri F, Cannella M, Giallonardo P, Maglione V,

as genetic modifiers of Huntington disease. Trends
Genet 23, 531–533.
12 Goehler H, Lalowski M, Stelzl U, Waelter S, Stroedicke
M, Worm U, Droege A, Lindenberg KS, Knoblich M,
Haenig C et al. (2004) A protein interaction network
links GIT1, an enhancer of huntingtin aggregation, to
Huntington’s disease. Mol Cell 15, 853–865.
13 Kaltenbach LS, Romero E, Becklin RR, Chettier R,
Bell R, Phansalkar A, Strand A, Torcassi C, Savage J,
Hurlburt A et al. (2007) Huntingtin interacting proteins
are genetic modifiers of neurodegeneration. PLoS Gene
3, 689–708.
14 Hyun TS & Ross TS (2004) HIP1: trafficking roles and
regulation of tumorigenesis. Trends Mol Med 10, 194–
199.
15 Kalchman MA, Koide HB, McCutcheon K, Graham
RK, Nichol K, Nishiyama K, Kazemi-Esfarjani P,
Lynn FC, Wellington C, Metzler M et al. (1997) HIP1,
a human homologue of S. cerevisiae Sla2p, interacts
with membrane-associated huntingtin in the brain. Nat
Genet 16, 44–53.
16 Wanker EE, Rovira C, Scherzinger E, Hasenbank R,
Wa
¨
lter S, Tait D, Colicelli J & Lehrach H (1997) HIP-
I: a huntingtin interacting protein isolated by the yeast
two-hybrid system. Hum Mol Genet 6, 487–495.
17 Hackam AS, Yassa AS, Singaraja R, Metzler M, Gut-
ekunst CA, Gan L, Warby S, Wellington CL, Vaillan-
court J, Chen N et al. (2000) Huntingtin interacting

llancourt JP, Houtzager V et al. (2002) Recruitment
and activation of caspase8 by the Huntingtin-interacting
protein HIP1 and a novel partner Hippi. Nat Cell Biol
4, 95–105.
24 Majumder P, Chattopadhyay B, Mazumder A, Das P &
Bhattacharyya NP (2006) Induction of apoptosis in cells
expressing exogenous Hippi, a molecular partner of
huntingtin-interacting protein Hip1. Neurobiol Dis 22,
242–256.
25 Rao DS, Hyun TS, Kumar PD, Mizukami IF, Rubin
MA, Lucas PC, Sanda MG & Ross TS (2002) Hunting-
tin-interacting protein 1 is overexpressed in prostate
and colon cancer and is critical for cellular survival.
J Clin Invest 110, 351–360.
26 Rao DS, Chang JC, Kumar PD, Mizukami I, Smith-
son GM, Bradley SV, Parlow AF & Ross TS (2001)
Huntingtin interacting protein 1 is a clathrin coat
binding protein required for differentiation of late
spermatogenic progenitors. Mol Cell Biol 21, 7796–
7806.
27 Bradley SV, Hyun TS, Oravecz-Wilson KI, Li L, Wald-
orff EI, Ermilov AN, Goldstein SA, Zhang CX, Drubin
DG, Varela K et al. (2007) Degenerative phenotypes
caused by the combined deficiency of murine HIP1 and
HIP1r are rescued by human HIP1. Hum Mol Genet 16,
1279–1292.
28 Khatchadourian K, Smith CE, Metzler M, Gregory
M, Hayden MR, Cyr DG & Hermo L (2007) Struc-
tural abnormalities in spermatids together with
reduced sperm counts and motility underlie the repro-

Apoptosis 13, 437–447.
35 Cheng CM, Huang SP, Chang YF, Chung WY & Yuo
CY (2003) The viral death protein Apoptin interacts
with Hippi, the protein interactor of Huntingtin-inter-
acting protein 1. Biochem Biophys Res Commun 305,
359–364.
36 Houde C, Dickinson RJ, Houtzager VM, Cullum R,
Montpetit R, Metzler M, Simpson EM, Roy S, Hayden
MR, Hoodless PA et al. (2006) Hippi is essential for
node cilia assembly and Sonic hedgehog signaling. Dev
Biol 300, 523–533.
37 Mao J, Maye P, Kogerman P, Tejedor FJ, Toftgard R,
Xie W, Wu G & Wu D (2002) Regulation of Gli1 tran-
scriptional activity in the nucleus by Dyrk1. J Biol
Chem 277, 35156–35161.
38 Mills IG, Gaughan L, Robson C, Ross T, McCracken
S, Kelly J & Neal DE (2005) Huntingtin interacting
protein 1 modulates the transcriptional activity of
nuclear hormone receptors. J Cell Biol 70, 191–200.
39 Majumder P, Chattopadhyay B, Sukanya S, Ray T,
Banerjee M, Mukhopadhyay D & Bhattacharyya NP
(2007) Interaction of HIPPI with putative promoter
sequence of caspase-1 in vitro and in vivo. Biochem
Biophys Res Commun 353, 80–85.
40 Majumder P, Choudhury A, Banerjee M, Lahiri A &
Bhattacharyya NP (2007) Interactions of HIPPI, a
molecular partner of Huntingtin interacting protein
HIP1, with the specific motif present at the putative
promoter sequence of the caspase-1, caspase-8 and
caspase-10 genes. FEBS J 274, 3886–3899.

activations and decreased mitochondrial complex II
activity in cells expressing exogenous huntingtin exon 1
containing CAG repeat in the pathogenic range. Cell
Mol Neurobiol 27, 1127–1145.
47 Ona VO, Li M, Vonsattel JP, Andrews LJ, Khan SQ,
Chung WM, Frey AS, Menon AS, Li XJ, Stieg PE
et al. (1999) Inhibition of caspase1 slows disease pro-
gression in a mouse model of Huntington’s disease.
Nature 399, 263–267.
48 Vis JC, Schipper E, de Boer-van Huizen RT, Verbeek
MmM, de Waal RM, Wesseling P, Ten Donkelaar HJ
& Kremer B (2005) Expression pattern of apoptosis-
related markers in Huntington’s disease. Acta Neuropa-
thol (Berl) 109, 321–328.
N. P. Bhattacharyya et al. HIP-1 & HIPPI mediated apoptosis and transcription
FEBS Journal 275 (2008) 4271–4279 ª 2008 The Authors Journal compilation ª 2008 FEBS 4279