KIF1Bβ Functions as a Haploinsufficient Tumor Suppressor Gene Mapped to Chromosome 1p36.2 by Inducing Apoptotic Cell Death*

Deletion of the distal region of chromosome 1 frequently occurs in a variety of human cancers, including aggressive neuroblastoma. Previously, we have identified a 500-kb homozygously deleted region at chromosome 1p36.2 harboring at least six genes in a neuroblastoma-derived cell line NB1/C201. Among them, only KIF1Bβ, a member of the kinesin superfamily proteins, induced apoptotic cell death. These results prompted us to address whether KIF1Bβ could be a tumor suppressor gene mapped to chromosome 1p36 in neuroblastoma. Hemizygous deletion of KIF1Bβ in primary neuroblastomas was significantly correlated with advanced stages (p = 0.0013) and MYCN amplification (p < 0.001), whereas the mutation rate of the KIF1Bβ gene was infrequent. Although KIF1Bβ allelic loss was significantly associated with a decrease in KIF1Bβ mRNA levels, its promoter region was not hypermethylated. Additionally, expression of KIF1Bβ was markedly down-regulated in advanced stages of tumors (p < 0.001). Enforced expression of KIF1Bβ resulted in an induction of apoptotic cell death in association with an increase in the number of cells entered into the G2/M phase of the cell cycle, whereas its knockdown by either short interfering RNA or by a genetic suppressor element led to an accelerated cell proliferation or enhanced tumor formation in nude mice, respectively. Furthermore, we demonstrated that the rod region unique to KIF1Bβ is critical for the induction of apoptotic cell death in a p53-independent manner. Thus, KIF1Bβ may act as a haploinsufficient tumor suppressor, and its allelic loss may be involved in the pathogenesis of neuroblastoma and other cancers.

Neuroblastoma is one of the most common malignant solid tumors occurring in infancy and childhood and accounts for 10% of all pediatric cancers (1). Neuroblastomas are derived from sympathetic neuroblasts with various clinical outcomes from spontaneous regression because of neuronal differentiation and/or apoptotic cell death to malignant progression. Extensive cytogenetic and molecular genetic studies identified that genetic abnormalities such as loss of short arm of chromosome 1 (1p), amplification of MYCN, and 17q gain are frequently observed and often associated with poor clinical outcome (2,3). The actual prevalence of 1p deletion in neuroblastoma is ϳ35% (4 -9). The deleted regions were extensively mapped to identify the candidate tumor suppressor gene(s) that has been deleted out from this region (10 -17). A chromosomal locus 1p36 is frequently deleted in aggressive neuroblastoma, pheochromocytoma, colon, liver, brain, breast, and other cancers (18,19). Transfer of 1p chromosome segments into neuroblastoma-derived cell line NGP.1A.TR1 resulted in a significant suppression of their tumor formation (20). Furthermore, previous studies indicated that there is no single site of deletion on the distal part of 1p36, but there are at least three discrete regions that are commonly deleted in neuroblastoma, indicating that they may harbor potential tumor suppressor gene(s) (8).
Tumor suppressor genes, one of the main classes of cancerassociated genes, encode inhibitors of cell proliferation and/or activators of apoptotic cell death and are involved in a variety of molecular mechanisms behind cell growth suppression (21). Tumor suppressor genes frequently mutated in other malignancies do not appear to play a major role in the generation of neuroblastoma, indicating that development of this type of tumor employs one or more previously unidentified genetic pathways. To date, a majority of candidate tumor suppressor genes has been identified by mapping the minimal deleted region and searching for the intact homologous region of mutated genes. This experimental strategy fails when the second allele is silenced by promoter hypermethylation or the targeted gene is haploinsufficient for tumor suppression, a situation where loss of one allele confers a selective advantage for tumor growth. Several examples of such haploinsufficiency for tumor suppression have been demonstrated in the case of p27 KIP1 , p53, and PTEN (22,23).
We and other investigators have previously identified a 500-kb homozygous deletion at 1p36.2 harboring at least six genes, PEX14, UFD2a, KIF1B, CORT, DFF45, and PGD, in a neuroblastoma-derived cell line NB1/C201 (12,15,24). In this study, we have demonstrated that only KIF1B␤, a member of the kinesin 3 family genes (25), might be a tumor suppressor gene mapped to chromosome 1p36 in neuroblastoma. Kinesins are microtubule-dependent intracellular motors involved in the transport of organelles, vesicles, protein complexes, and RNA to specific destinations (26,27). KIF1B encodes two alternatively spliced isoforms, including KIF1B␣ and KIF1B␤, and both form homodimers and transport mitochondria and synaptic vesicle precursors, respectively (28). The NH 2 -terminal motor domain of KIF1B␣ is identical to KIF1B␤, whereas COOH-terminal tails share no structural homology. A point mutation in the ATP-binding site within the motor domain of KIF1B␤ has been closely linked to Charcot-Marie Tooth disease type 2A (29).
In this study, we cloned a full-length KIF1B␤ cDNA, generated recombinant adenovirus encoding KIF1B␤, and examined its biological role in neuroblastoma and other cell lines. We systematically analyzed KIF1B␤ for LOH, 2 mutation, and promoter methylation. Our genetic and functional analyses clearly showed that KIF1B␤ is a tumor suppressor, although not a classic one. KIF1B␤ might act as a haploinsufficient tumor suppressor, and its down-regulation might potentially contribute to tumorigenesis of cancers, including neuroblastoma.

EXPERIMENTAL PROCEDURES
Cell Lines and Tumor Samples-Human neuroblastoma (NB) cell lines such as SH-SY5Y, NB1, and SK-N-BE were grown in RPMI 1640 medium supplemented with heat-inactivated fetal bovine serum, 100 units/ml penicillin, and 100 g/ml streptomycin. NMuMG, COS7, HEK293, and HeLa cells were grown in Dulbecco's modified Eagle's medium containing 10% heat-inactivated fetal bovine serum, 100 units/ml penicillin, and 100 g/ml streptomycin. Tumor DNA and RNA samples were obtained from our Neuroblastoma Resource Bank. Informed consent was obtained at each hospital.
GSE-mediated Tumor Formation in Nude Mice-GSE assay was performed as described previously (30). In brief, a cDNA fragment (nucleotide number 2658 -3115 of GenBank TM accession number AB017133) corresponding to the unique region of KIF1B␤ was amplified by PCR-based strategy and subcloned into the HpaI site of the pLXSN vector in an antisense orientation to give pLXSN-antisense KIF1B␤. NMuMG mammary gland cells (1 ϫ 10 6 cells) infected with pLXSN or with pLXSN-antisense KIF1B␤ were inoculated subcutaneously into the femoral region of nude mice. In the experiments using live animals, we strictly followed the Chiba Cancer Center Research Institute guidelines and protocols for handling live animals.
Construction of Expression Plasmids and Recombinant Adenovirus-KIF1B␤ splicing variants I, III, and IV fused to the FLAG epitope at their NH 2 termini were amplified by PCR using cDNA prepared from CHP134 cells as a template and subcloned into pcDNA3.1 (Invitrogen). KIF1B␤-GFP deletion constructs were produced by PCR-based amplification. The recombinant adenovirus was constructed as described previously (31). Briefly, KIF1B␤ cDNA was subcloned into pHMCMV6 adeno-shuttle vector. The shuttle vector was then digested with I-Ceu I and PI-Sce I and inserted into the identical restriction sites of the adenovirus expression vector pAdHM4. All of the recombinant vectors were verified by DNA sequencing. Recombinant adenoviruses were produced by transfecting the PacI-digested expression constructs into HEK293 cells. An expression vector encoding GFP was used to monitor the efficiency of infection.
Mutation Analysis-For the detection of KIF1B␤ mutations, we designed primer sets covering the motor domain and 2 kb of the 5Ј-upstream region of KIF1B␤. After PCR-based amplification, PCR products were separated by 5% nondenaturing polyacrylamide gels. After electrophoresis, PCR products were gelpurified and subcloned into pGEM-T Easy Vector (Promega), and their DNA sequences were determined by an automated DNA sequencer (Applied Biosystems).
Flow Cytometry-Cells were fixed in ice-cold 70% ethanol, treated with 50 mM sodium citrate, 100 g/ml RNase A, 50 g/ml propidium iodide and subjected to FACS analysis (BD Biosciences) according to the manufacturer's instructions.
Construction of KIF1B␤ siRNA Expression Vector-An siRNA expression vector termed pMuniH1, in which the cytomegalovirus promoter of pcDNA 3.1 was replaced with the H1 promoter, was generated. Sense and antisense oligonucleotides for KIF1B␤ (nucleotide number 371-389 of GenBank TM accession number AB017183) were joined by a 9-base loop, annealed, and subcloned into pMuniH1.
Methylation-specific PCR-The methylation status of the promoter region of KIF1B␤ was assessed by methylation-specific PCR as described previously (32).
Cell Cycle Analysis-Cells were fixed in 3.7% formaldehyde and permeabilized with 0.2% Triton X-100 and DNA was stained with 0.1 g/ml of 4Ј,6Ј-diamidino-2-phenylindole. Cellular DNA content was analyzed by laser scanning cytometry (LSC2 System, Olympus).
Array-CGH Analysis-Array CGH analysis of 112 sporadic primary neuroblastomas using a chip carrying 2,464 bacterial artificial chromosome clones was conducted as described previously (33). All array-CGH data are available at NCBI Gene Expression Omnibus (GEO, www.ncbi.nlm.nih.gov) with accession number GSE 5784.
Caspase Assay-Caspase activity was measured by using caspase-3/7 assay system (Promega) according to the manufacturer's instructions.
Statistics-The Student's t test was used as a statistical method. Statistical significance was declared if the p value was Ͻ0.05.

Identification of KIF1B␤ as a Candidate Tumor Suppressor Mapped to
Chromosome 1p36.2-To search for a candidate tumor suppressor gene(s), we first transferred each of the above-mentioned six genes into NB1 and nontransformed NMuMG mouse epithelial cells (30), and we found that only KIF1B␤ induces growth suppression in a dosedependent manner (Fig. 1A). In contrast, our preliminary observations indicated that its alternative splicing variant KIF1B␣ lacking a COOH-terminal rod region has marginal effects on cell growth in NB1 cells (data not shown). In support of these results, siRNA-mediated knockdown of KIF1B␤ in HeLa cells without 1p loss markedly enhanced their cell growth (Fig. 1B). In addition, enforced expression of KIF1B␤ induced growth retardation in p53-deficient H1299 cells and HeLa cells in which p53 is inactivated because of the presence of E6-AP (data not shown).
Overexpression of KIF1B␤ in Neuroblastomas-induced Apoptotic Cell Death-During PCR-based screening of human KIF1B␤ cDNA from human neuroblastoma cell lines, we identified at least four splicing variants that lacked exons 14 and/or 15 (supplemental Fig. S1). Similar splicing variants have also been observed in mice and rats (34). We successfully generated recombinant adenoviruses for human KIF1B␤-I, -III, and -IV variants ( Fig. 2A). Enforced expression of these splicing variants promoted apoptotic cell death in both SH-SY5Y (without 1p loss) and NB1 neuroblastoma cell lines as determined by MTT and FACS analyses ( Fig. 2A and supplemental Fig. S2). Consistent with these results, colony formation assay showed that all these KIF1B␤ splicing variants strongly reduced the number of drug-resistant colonies in SH-SY5Y and NB1 neuroblastoma cells (Fig. 2B). These findings suggest that multiple KIF1B␤ splicing isoforms  . Twenty four hours after infection, infected SH-SY5Y and NB1 cells were seeded at a density of 1 ϫ 10 3 cells/96-well plates and allowed to attach. Ninety six hours after infection, 10 l of MTT solution was added to each well and incubated for 3 h at 37°C (left panel). Right panels show the expression of the indicated splicing variants of KIF1B␤ as examined by immunoblotting (IB) with anti-FLAG antibody. B, colony formation assay. SH-SY5Y and NB1 cells were transfected with an empty plasmid or with the indicated expression plasmids. Forty eight hours after transfection, cells were transferred into fresh medium containing 500 g/ml of G418 and incubated for 2 weeks. After selection with G418, G418-resistant viable colonies were stained with Giemsa solution, and number of colonies was scored. possess tumor suppressor activity. Intriguingly, the expression pattern of KIF1B␤ splicing variants was varied among various human tissues (supplemental Fig. S3).

Knockdown of KIF1B␤ Expression Accelerates Growth of NMuMG Cells and Tumor Formation in Nude
Mice-We then asked whether genetic disruption of KIF1B␤ gene could be critical for tumorigenesis. For this purpose, we employed a genetic suppressor element (GSE) strategy (30). A mouse genomic DNA corresponding to a KIF1B␤ cDNA fragment (nucleotide number 2658 -3115 of GenBank TM accession number AB017133) encoding the unique region of KIF1B␤ was subcloned into the retrovirus pLXSN vector in an antisense orientation to give pLXSN-KIF1B␤-AS. NMuMG cells, immortalized mouse mammary gland cells, stably infected with pLXSN-KIF1B␤-AS, showed more than 80% reduction in endogenous KIF1B␤ expression (Fig. 3A) and formed significantly larger colonies than empty vector-infected control cells in soft agar medium (Fig. 3B). In addition, all eight mice subcutaneously transplanted with NMuMG cells stably infected with pLXSN-KIF1B␤-AS displayed remarkable tumor growth (Fig.  3C). On the other hand, only two of eight mice transplanted with the empty vector-infected cells formed tumors, which were smaller in both cases (note log scale in Fig. 3C). The tumors formed by cells lacking KIF1B␤ expression were histologically diagnosed as poorly differentiated invasive ductal carcinoma and frequently metastasized to the lung (Fig. 4). Thus, it is likely that KIF1B␤ exerts tumor-suppressive function in vivo.
LOH of KIF1B␤ Locus Is Frequently Observed in Primary Advanced Neuroblastomas-We next sought to search for LOH at chromosome 1p36 in 112 sporadic neuroblastomas using array-based comparative genomic hybridization (array-CGH). Similar to previous reports, the smallest region of overlap at the distal region of chromosome 1p identified in 37 primary neuroblastomas with 1p loss was between 1p36.22 and 1pter and included KIF1B, CHD5, TP73, and SKI (supplemental Fig. S4). Thirty two percent (30/95) of neuroblastomas examined had lost one KIF1B␤ allele as determined by quantitative real time genomic PCR (Table 1). KIF1B␤ was hemizygously deleted in 18% of early neuroblastomas (stages 1 and 2, n ϭ 51), in 55% of advanced neuroblastomas (stages 3 and 4, n ϭ 38) (p ϭ 0.0013), in 13% of primary neuroblastomas with a single copy of MYCN (n ϭ 70), and in 84% of MYCN-amplified primary neuroblastomas (n ϭ 25) (p Ͻ 0.001). No homozygous deletion was detected in the primary tumors examined.   SEPTEMBER 5, 2008 • VOLUME 283 • NUMBER 36

JOURNAL OF BIOLOGICAL CHEMISTRY 24429
Decreased Expression of KIF1B␤ Is Associated with Monoallelic Loss of the Gene in Primary Neuroblastomas-We examined expression levels of KIF1B␤ mRNA in 102 primary neuroblastomas by using both semi-quantitative and quantitative real time PCRs. As shown in Fig. 5, A and B, expression levels of KIF1B␤ mRNA were significantly higher in tumors at favorable stages (1, 2, and 4s, 1.654 Ϯ 0.257, mean Ϯ S.E., n ϭ 60) than in those at advanced stages (3 and 4, 0.503 Ϯ 0.180, n ϭ 42, p Ͻ 0.001). To address whether its expression levels could be correlated with number of alleles at the KIF1B␤ gene locus, we examined primary tumors with a diploid karyotype. As shown in the lower panel of Fig. 5B, tumors with monoallelic loss of KIF1B␤ gene locus expressed significantly lower levels of KIF1B␤ mRNA (0.126 Ϯ 0.092, n ϭ 13) as compared with those with two KIF1B␤ alleles (0.364 Ϯ 0.035, n ϭ 16, p ϭ 0.019). These results suggest that KIF1B␤ is a haploinsufficient tumor suppressor gene in high risk neuroblastomas.
No Promoter Methylation and Rare Mutations Are Observed in Neuroblastoma Cell Lines and Primary Neuroblastomas-Our initial mutation searches of KIF1B␤ gene were focused on its motor domain and the proximal (ϳ2 kb) promoter region in 21 primary neuroblastomas with 1p36 LOH and in 17 neuroblastoma cell lines. As shown in Table 2, we identified only a silent mutation GCC-GCG (at codon 95) in two primary tumors, a 2-bp (CC) deletion (at Ϫ113/Ϫ114) and G-A base change (at Ϫ336) in the KIF1B␤ promoter region in three primary tumors and four neuroblastoma cell lines. Because these aberrations were also found in the control samples, it is likely that these base changes reflect single nucleotide polymorphisms of the Japanese population.
To further extend our mutation searches, we have examined the presence or absence of KIF1B␤ mutations within its whole coding region in 100 primary neuroblastoma tissues. Finally, we found out the missense mutations (N737S) in six independent cases. However, their functional significances remained unclear.
Methylation of CpG islands in the promoters has been considered to be another well recognized molecular mechanism behind the inactivation of the tumor suppressor gene. To determine whether the methylation of CpG island could contribute to the inactivation of KIF1B␤, the region spanning exon 1 and 5Ј-upstream sequences (nucleotide number Ϫ877 to ϩ106) of KIF1B␤ was cloned and analyzed for promoter activity by luciferase reporter assay. As shown in Fig. 6, A and B, KIF1B␤ promoter region existed at nucleotide position between Ϫ630 and Ϫ294. We then identified KIF1B␤ CpG islands within the promoter region and investigated whether these CpGs could be methylated in primary neuroblastomas as well as cell lines. The methylation-specific PCR analysis demonstrated that all of the CpG clusters are unmethylated, suggesting that KIF1B␤ is not inactivated by methylation (Fig. 6C).

The COOH-terminal Region between FHA and Pleckstrin Homology Domains of KIF1B␤ Is Responsible to
Induce Apoptotic Cell Death-To map a critical domain(s) of KIF1B␤ responsible for its tumor-suppressive function, we generated NH 2 -

KIF1B␤ Is a Haploinsufficient Tumor Suppressor
terminal deletion mutants of KIF1B␤ splicing isoform IV lacking motor domain (KIF1B␤-IV-Del 1-GFP), motor and FHA domains (KIF1B␤-IV-Del 2-GFP), coiled-coil domain (KIF1B␤-IV-Del 3-GFP), and FHA and coiled-coil domains (KIF1B␤-IV-Del 4-GFP) fused with enhanced green fluorescent protein at their COOH termini (Fig. 7A). COS7 cells were transfected with wild-type or with KIF1B␤-IV-Del-GFP fusion constructs and followed by live confocal laser scanning microscopy. Forty eight hours after transfection, KIF1B␤-GFP-positive cells began to lose their normal cell morphology. Seventy two hours after transfection, most of the cells underwent apoptotic cell death and detached from the cell culture dish (Fig. 7, B and D). Expression of splicing variants-I, -III, and -IV along with deletion mutants of splicing variant-IV lacking the motor  domain (with or without FHA domain) also induced apoptotic cell death. In contrast, expression of KIF1B␤ mutants lacking the COOH-terminal rod domain did not promote apoptotic cell death (Fig. 7, A and B). Under our experimental conditions, enforced expression of KIF1B␤ variant-IV resulted in a significant increase in the caspase activities (Fig. 7C), suggesting that KIF1B␤-mediated apoptotic cell death might be regulated in a caspase-dependent manner. Our further analysis using other deletion mutants revealed that the 807 amino acids death-inducing region is located between FHA and pleckstrin homology domains (data not shown).
To determine whether the kinesin activity of KIF1B␤ could be necessary for its tumor-suppressive function, we introduced a Q98L mutation within a consensus ATP-binding site of KIF1B␤-IV splicing variant (Fig. 8A). This mutation disrupts the motor function of KIF1B␤ (29). In addition, a KIF1B␤-IV splicing variant carrying two point mutations (Q560A and D568A) within its highly conserved amino acid residues of FHA domain, which may be critical for binding to Ser/Thr-phosphorylated motifs of the interacting proteins, was also generated. In addition to these two mutants, we also generated an additional mutant bearing Q98L, Q560A, and D568A. These three mutants, however, retained an ability to induce apoptotic cell death, suggesting that KIF1B␤-mediated apoptotic cell death does not require its ability to transport cargo using its motor domain (Fig. 8B).

DISCUSSION
In this study, we have shown that the KIF1B␤ gene is hemizygously deleted especially in aggressive primary neuroblastoma tumors, and its mutation is infrequent. The expression of KIF1B␤ was kept at quite a low level in aggressive neuroblastoma subsets, even though no methylation of its promoter region was observed. One of the well known haploinsufficient tumor suppressors is the cyclin-dependent kinase inhibitor p27 KIP1 (35). The heterozygous mice of p27 KIP1 developed tumors when mice were treated with tumor-promoting agents, and tumors retained the normal p27 KIP1 allele. Additionally, hemizygous loss of p27 KIP1 and/or reduced expression level of p27 KIP1 conferred poor prognosis in human cancers (36). Taken together, our present results suggest that, like p27 KIP1 , KIF1B␤ is a haploinsufficient tumor suppressor gene of neuroblastoma, and its function to induce apoptotic cell death is regulated in a p53-independent manner. Although homozygous deletion or mutations of KIF1B␤ were rarely detectable in this study, several losses of function mutations in the coding region of KIF1B␤ gene in a large number of primary neuroblastomas, pheochromocytomas, and medulloblastomas have now been identified. 3 Homozygous deletion of KIF1B in mice resulted in death just after birth because of apnea. However, heterozygous mice are viable with a phenotype resembling Charcot-Marie-Tooth disease type 2A (29). To date, no information has been available in the literature regarding spontaneous tumor formation in KIF1B-heterozygous mice. It is possible that these mice have not been followed long enough or that loss of one KIF1B allele is not sufficient for tumor formation and requires cooperating mutations for spontaneous tumor formation. Since there is functional disruption of wild-type p53 because of its mislocalization, haploinsufficiency of the KIF1B␤ gene might contribute to tumorigenesis of aggressive neuroblastomas with 1p LOH and MYCN amplification (37). KIF1B␤ might also be involved in tumorigenesis in combination with other contiguous 1p36.3 genes such as p73 (38) and CHD5 (39).
Finally, we have identified four different splicing variants of KIF1B␤. However, colony formation assay revealed that all of the splicing variants almost equally suppress cell growth, indicating that its tumor-suppressive function may not be dependent on alternative splicing events. The deletion construct termed Del 2-GFP encoding amino acid residues 637-1576 3 S. Schlisio and W. G. Kaelin, Jr., personal communication. induced apoptotic cell death similar to wild-type KIF1B␤. Therefore, this region containing two predicted coiled-coils (amino acid residues 668 -737 and 841-863) alone is sufficient for pro-apoptotic function of KIF1B␤. The coiled-coil motifs are amphipathic oligomerization motifs. The tumor suppressor par-4 with a potential coiled-coil structure induced apoptotic cell death in prostate cancer cell lines (40,41). Moreover, a putative coiled-coil domain of potential tumor suppressor protein, Prohibitin, has been shown to be sufficient to repress E2F1-mediated transcription and induction of apoptotic cell death (42).
In neuroblastoma, polyploidy is very common, which is often associated with a better prognosis. The precise molecular mechanisms underlying this phenomenon still remain unclear. Recently, defects in mitotic spindle check point gene products such as MAD1, MAD2, BUB1, BUB3, and BUBR1 have been implicated in the generation of polyploidy (43). Intriguingly, attached cells expressing GFP-tagged KIF1B␤ splicing variants exhibited a perturbation of G 2 /M progression and multinucleation (supplemental Fig. S5). The precise molecular mechanisms by which KIF1B␤ could promote these cellular abnormalities and apoptotic cell death are currently unknown. On the other hand, down-regulation of KIF1B␤ resulted in augmented cell proliferation in vitro and tumor formation in vivo, indicating that KIF1B␤ might have a critical role in the regulation of mitosis like other mitotic kinesins (44). It is conceivable that KIF1B␤ might act in a dominant inhibitory manner to sequester fundamental cytoplasmic factors that are required for proper cell cycle progression. In this connection, we are undertaking to identify the KIF1B␤-binding partner(s), which might clarify the molecular mechanisms behind growth suppression and/or apoptotic cell death mediated by KIF1B␤.
The nerve growth factor (NGF) dependence of tumor cells through the TrkA-p75 NTR receptor complex plays a critical role in the regulation of the spontaneous regression and differentiation in neuroblastoma (45). NGF depletion-induced apoptotic cell death is blocked in aggressive neuroblastoma (46). The findings showing that expression of KIF1B␤ also increases during apoptotic cell death triggered by NGF depletion in PC12 cells 3 strengthen the significance of the tumor suppressor function of KIF1B␤ in primary neuroblastomas and pheochromocytoma. Indeed, some kinesin family proteins are involved in the regulation of apoptotic cell death in developing neurons (47). In conclusion, our present results unveiled that KIF1B␤, mapped to chromosome 1p36.2, is the candidate tumor suppressor gene of the kinesin family functioning in a manner of haploinsufficiency.