Despite the importance of understanding the normal function of Htt for both basic biology and for HD, its function remains largely unknown [9, 34]. The generation of Hdh-null MEF cell lines described in our study provides a new and unbiased approach to search for novel Htt functions.
The Htt directly binds to the InsP3R, an intracellular Ca2+ release channel [23, 24]. The expression of mutated Htt has been shown to affect the InsP3R activity  and mitochondrial Ca2+ signals and bioenergetics [25–30]. Thus, in the first series of experiments we evaluated InsP3R-mediated Ca2+ cytosolic and mitochondrial Ca2+ signals in Hdh-null MEF cells. As a result of these experiments we found that InsP3R sensitivity to stimulation by InsP3 was reduced in the absence of Hdh (Figs 3 and 4). We further found that Htt appears to be dispensable for ER-mitochondrial Ca2+ coupling (data not shown). Thus the altered InsP3R-induced cytoplasmic and mitochondrial calcium signaling in the Hdh-null MEF cells may result from the lack of Hdh by itself and does not necessarily require a secondary change in gene regulation.
Interestingly, a large number of Ca2+-related genes, such as CACNA2D3 (calcium channel, voltage-dependent, alpha2/delta subunit 3), ITPR1 (inositol 1,4,5-trisphosphate receptor 1), HOMER1 (homer homolog 1), ATP2A2 (ATPase, Ca2+ transporting, cardiac muscle, slow twitch 2), DRD2 (dopamine receptor 2), PRKCB1 (protein kinase C beta 1), PDE1B (phosphodiesterase 1B, Ca2+-calmodulin dependent), ATP2B2 (ATPase, Ca2+ transporting, plasma membrane 2), CAMK2B (calcium/calmodulin-dependent protein kinase II, beta), PLCB1 (phospholipase C, beta 1), RGS4 (regulator of G-protein signaling 4), and CAMK2A (calcium/calmodulin-dependent protein kinase II alpha), have been recently reported to be consistently and significantly downregulated in a striatal region of symptomatic human HD patients and aging HD mouse models . These results are in agreement with the "Ca2+ hypothesis of HD"  and with a direct role of Htt in intracellular Ca2+ signaling supported by our experiments.
Many studies suggested that Htt also plays a major role in control of gene transcription [33, 34]. To search for changes in gene transcription resulting from deletion of Htt gene, we performed a genome-wide comparison of transcription profiles in MEF cells expressing a single copy of Hdh (Hdh-HET cells) and in MEF cells which lack Hdh expression (Hdh-KO cells). To minimize sources of variability, the Hdh-HET and Hdh-KO MEF cells were generated in parallel experiments and on identical genetic background. From our annotation analysis, we found that a large group of affected genes play a role in embryonic development (Additional file 1). This result was not unexpected because Htt is essential for embryonic development, and complete inactivation of Htt expression in knock-out mice causes early embryonic lethality [10–12]. The functions of these genes may provide additional clues about the mechanism responsible for embryonic lethality in Hdh knockout mice, for example there are some similar phenotypic manifestations between Hdh nullizygous embryos and knockout mutants lacking fibroblast growth factor receptor1(fgfr1) . Interestingly, we found fgfrl1 message is downregulated approximately 2-fold in Hdh-KO MEF cells when compared to Hdh-HET cells (Additional files 1 and 2).
After gastrulation, Htt is important for neurogenesis – mice carrying a <50% dose of wild-type Htt display profound malformations of the cortex and striatum . The neuronal inactivation of Htt during mid- to late gestation, for example, leads to neurological abnormalities and progressive degeneration (apoptotic cells in the hippocampus, cortex and striatum, and a lack of axons) . Our analysis revealed a number of genes involved in nervous system development and function which were affected in Hdh-KO MEF cell lines (Additional file 1), and this list should also provide useful information to guide further studies of Htt's normal function in the nervous system. For example, Sox-2 expression was absent from Hdh-KO MEFs. Sox (Sry-related HMG box) genes encode transcription factors regulating crucial developmental decisions in different systems. Sox2 is expressed in, and is essential for, totipotent inner cell mass stem cells and other early multipotent cell lineages, and its ablation causes early embryonic lethality . In many different species, Sox2 is a marker of the nervous system from the beginning of its development, it maintains a stem-cell like state and actively inhibits neuronal differentiation, Sox2 deficiency causes neurodegeneration and impaired neurogenesis in the adult mouse brain. Does the absence of Sox-2 play some role in the early embryonic lethality and neurodegeneration in Hdh knock-out mice and conditional knock-out mice respectively? Further studies are required to answer these questions. Sox-11 is another sox family gene which changes dramatically in Hdh-KO MEF cell lines (increases about 10-fold, see Additional files 1 and 2). The widespread expression of sox-11 in both the central and peripheral nervous system suggests that sox-11 plays a general role in neuronal development, and its changes in Hdh-KO cells merit further investigation.
Another group of genes whose expression was significantly affected in Hdh-KO MEF cells are the genes related to lipid metabolism. It has been reported in another microarray analysis using clonal striata-derived cells, that genes involved in lipid metabolism were affected after expressing different N-terminal 548-amino-acid Htt fragments . Moreover, recent biochemical data indicated that Htt binds to caveolin and plays a direct role in cholesterol metabolism . All these data suggested that Htt plays an important role in lipid metabolism, which may be affected by HD mutation. Indeed, RXRG (Retinoic acid receptor RXR-gamma) and RBP4 (retinol binding protein 4) are consistently downregulated in a striatal region of symptomatic human HD patients and aging HD mouse models .
From our analysis we also found calcium channel voltage-dependent alpha2/delta subunit 1 (Cacna2d1) was down-regulated about 2 fold (Additional files 1 and 2), interestingly the same CACNA2D1 protein has been recently identified as novel Htt-binding partner in unbiased mass-spectroscopy screen . A closely related alpha2/delta subunit 3 (Cacna2d3) was reported on the 3rd place on the list of the genes which are consistently and significantly downregulated in a striatal region of symptomatic human HD patients and aging HD mouse models . As discussed above, these results indicate that Htt may play a role in regulation of Ca2+ channel activity and Ca2+ signaling in cells, consistent with Ca2+ hypothesis of HD [22, 43].